Báo cáo khoa học: Invited speakers pot

Invited speakers
I01
Please view Plenary lectures
I02
Abstract missing, please view Abstract Addendum
I03
Aquaporin water channels in health and
disease
M. Amiry-Moghaddam
University of Oslo, Oslo, Norway
The water permeability of biological membranes has been a long-
standing problem in physiology, but the proteins responsible for
this remained unknown until discovery of the aquaporin 1
(AQP1) water channel protein. Peter Agre received Nobel prize in
Chemistry in 2003 for this discovery. AQP1 is selectively perme-
ated by water driven by osmotic gradients. The atomic structure
of human AQP1 has been defined. Each subunit of the tetramer
contains an individual aqueous pore that permits single-file
passage of water molecules but interrupts the hydrogen bonding
needed for passage of protons. At least 12 mammalian aquaporins
have been identified, and these are selectively permeated by water
(aquaporins) or water plus glycerol (aquaglyceroporins) and more
than 200 members of the aquaporin family have been found in
plants, microbials, invertebrates and vertebrates. Research in the
past decade has shown that aquaporins are not only involved in
important physiological processes such as maintenance of body
water homeostasis, but are also involved in pathological condi-
tions such as nephrogenic diabetes insipidus, cataract, metabolic
syndrome, brain edema and epilepsy.
I04
Abstract missing, please view Abstract Addendum

I05
Electron transfer routes in photosynthetic
membranes – impact on biohydrogen
production
E-M. Aro, P. Zhang, M. Eisenhut, Y. Allahverdiyeva
and N. Battchikova
University of Turku, Turku, Finland
Optimization of electron transfer from water to biohydrogen pro-
duction in a model organism Synechocystis sp. PCC 6083
involves several steps. PSII function can be improved by intro-
ducing a proper PSII reaction center D1 protein (encoded by the
psbA gene family). To this end, an expression of a specific psbA
gene that encodes a D1’ protein was detected under anaerobic
conditions. We have also demonstrated a novel and crucial func-
tion for Flavodiiron (FDP) proteins Flv2 and Flv4 in photopro-
tection of PSII. The rate of accumulation of flv2 and flv4
transcripts upon shift of cells from high to low CO
2
is strongly
dependent on light intensity. Characterization of FDP inactiva-
tion mutants revealed a specific decline in PSII centers and
impaired translation of the D1 protein in Dflv2 and Dflv4 when
grown at air level CO
2
whereas at high CO
2
the FPDs were dis-
pensable. Dflv2 and Dflv4 were also more susceptible to high light
induced inhibition of PSII than WT or Dflv1 and Dflv3. Of the
four flavodiiron proteins (Flv1-4) in Synechocystis 6803, a physi-

ological function of Flv1 and Flv3 is in the Mehler reaction. Up
to 30% of electrons derived from water by PSII may be directed
to molecular oxygen via Flv1 and Flv3, and thus this route might
seriously compete for electrons with the hydrogenase. Besides
FDPs, the multiple NDH-1 complexes in cyanobacterial thyla-
koid membranes have a crucial role in electron transfer reactions,
particularly in cyclic electron transfer around PSI, in respiratory
electron transfer and in carbon concentrating mechanisms. More-
over, interplay between the FDPs and NDH-1 complexes is dem-
onstrated to occur in electron transfer reactions.
I06
Global transcriptional regulation of the gut
microbiota and its impact on host physiology
F. Ba
¨
ckhed
University of Gothenburg, Gothenburg, Sweden
The adult intestine contains around 100 trillion bacteria, a num-
ber 10 times greater than the number of human cells in our body.
This complex ecosystem (gut microbiota) has complemented our
own genome with several functions that affects human health:
modulation of metabolism, development of the immune system,
and protection against enteric infections. Recent data have impli-
cated the gut microbiota to be involved in obesity, which is asso-
ciated with an altered gut microbiota. The gut microbiota can
affect host metabolism either directly or indirectly by affecting
gene expression. Germ-free mice have provided an important tool
to investigate the underlying molecular mechanisms for how the
gut microbiota affects host physiology. By using metabolomics
and lipidomics we have recently found that the gut microbiota

directly affects the serum metabolome and serum, liver, and adi-
pose lipidomes. By comparing the transcriptional profile along
the length of the gut in germ-free and conventionally raised mice
we have demonstrated altered expression of several hormones
that are produced by the epithelium. Additional experiments
revealed that microbial regulation of gene expression in the small
intestine could directly affect host metabolism.
Furthermore, we could demonstrate a rapid induction of genes
involved in the innate immune system in the epithelium as well
as recruitment of immune cells to the mucosa. Taken together
the gut microbiota should be considered as an organ by itself
that has major effects on host metabolism and physiology, were
perturbations may cause or promote disease.
I07
Folding and redox processes in the
mitochondria
L. Banci
CERM & Department of Chemistry, University of Florence, Sesto
Fiorentino (Florence), Italy
A number of proteins undergo their folding through various
states that are dependent on the cellular compartment and on
compartment-specific protein components. A number of nuclear-
encoded proteins, lacking the mitochondrial target sequence,
enter mitochondria through transmembrane channels and get
trapped in the IMS through oxidative folding processes. These
proteins are characterized by disulfide bonds whose formation is
combined to protein folding. Disulfide bond formation is kineti-
Invited speakers Abstracts
FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies 5
cally slow and is catalyzed by a protein, Mia40, which is part of

a disulfide relay system. In this system the electron flow goes
from Mia40 substrates up to cytochrome c and cytochrome c oxi-
dase. Significantly some of the Mia40 substrates are proteins
responsible for incorporating copper in cytochrome c oxidase.
This behaviour shows how various processes occurring in the
same cellular compartment are tightly connected and interlinked,
and therefore only a charactrization at ‘system’ level can fully
describe functional processes. A few examples of folding pro-
cesses will be presented and discussed.
I08
New insights into hepatitis C virus replication
and persistence
R. Bartenschlager
Department of Infectious Diseases, Molecular Virology, University
of Heidelberg, Heidelberg, Germany
Hepatitis C viruses (HCV) comprise the sole genus hepacivirus in
the family Flaviviridae. These viruses have in common a single
strand RNA genome of positive polarity encoding for a single
polyprotein that is cleaved by host and viral proteases. Studies of
the HCV replication cycle have become possible by the develop-
ment of highly efficient and robust cell culture systems. By using
these techniques four molecules (CD81, scavenger receptor class
B type I; claudin-1 and occludin) that are essential for infection
of hepatocytes have been identified and validated. Moreover,
important insights into the biogenesis and architecture of the
membranous replication complex induced upon viral infection
have been gained. Finally, it was found that HCV assembly
occurs in close association with lipid droplets and the host cell
machinery required for the synthesis of very-low-density-lipopro-
tein (VLDL). As a net result of the assembly reaction, infectious

virus particles are formed that are unique in structure and com-
position and that resemble most closely LDL.
A hallmark of HCV infection is the high rate of persistence
(~80%). It was found that the viral serine-type protease NS3 is a
key factor that blocks the induction of antiviral cytokines, esp.
type 1 interferon (IFN) by proteolytic cleavage of adaptor pro-
teins involved in RIG-I and TLR-3 dependent signalling. Apart
from blocking innate immunity, HCV overcomes adaptive
immune responses by multiple strategies including antigenic vari-
ability. Moreover, the tight association of HCV particles with lip-
ids appears to impair virus neutralization. Thus HCV utilizes
multiple strategies to establish persistence.
I09
The billion protein question
A. Bateman
Wellcome Trust Genome Campus, Hinxton, UK
Next generation sequencing is pouring out incredible amounts of
data. We are inevitably moving towards the day when we know
1 billion protein sequences. This is both exciting and terrifying. I
will discuss ways we might deal with the data deluge. Will we all
need to become Bioinformaticians?
I10
Salmonella metabolism during infection
D. Bumann
Biozentrum, University of Basel, Basel, Switzerland
Metabolism is an important aspect of Salmonella biology during
infection. However, individual metabolic perturbations rarely
diminish Salmonella virulence in a mouse typhoid fever model.
To understand underlying causes for this large-scale enzyme dis-
pensability, we used in silico modeling combined with systematic

experimental analysis. Our genome-scale metabolic in silico
model correctly predicts virulence phenotypes of more than 92%
of 849 experimentally characterized Salmonella mutants. Model-
ing and experimental analysis of multiple metabolic mutations
revealed a minor impact of network redundancy on overall
robustness. However, the host microenvironment provides diverse
nutrients that are mainly responsible for extensive Salmonella
robustness against perturbation. Interestingly, genome compari-
sons suggest that many other pathogens might encounter similar
nutritional patterns in their hosts that markedly differ from habi-
tats of environmental microbes.
I11
Designer cellulosomes: Synthetic
multi-enzyme macromolecular complexes
E. Bayer
Department of Biological Chemistry, The Weizmann Institute of
Science, Rehovot, Israel
Cellulosomes are intricate multi-enzyme machines produced by
anaerobic cellulolytic microorganisms, designed for efficient
decomposition of plant cell wall polysaccharides, notably cellu-
lose – the most abundant renewable organic polymer on Earth.
The cellulosome complex consists of interlocking, multi-modular,
structural and enzymatic subunits, which fit together in a Lego-
like arrangement. We have harnessed the molecular logic of the
cellulosome components and have developed methodologies to
re-tool its precise structural organization. For this purpose, func-
tional modular parts of cellulosome subunits from different
microorganisms are mixed and matched by recombinant means
to produce chimaeric products, which can be fitted together in a
controlled manner into novel cellulosome-like structures. The

resultant designer cellulosomes are functional, and their cellulo-
lytic capacities approach and frequently surpass those of equiva-
lent free enzyme systems. Our approach is designed to better
understand these intercomponent interactions, to discover how
the cellulosome is constructed and how cellulosome architecture
contributes to the enhanced synergistic activities of its enzyme
components. Knowledge of these interactions provides a broad
platform for biotechnological and nanotechnological applica-
tions, including prospects for conversion of plant cell wall bio-
mass to biofuels – a crucial goal of global importance in the 21st
century for all mankind.
I12
Pathophysiology of the mitochondrial
permeability transition
P. Bernardi
Biomedical Sciences, University of Padova, Padova, Italy
The mitochondrial permeability transition (PT) is a Ca
2+
-depen-
dent increase of mitochondrial inner membrane permeability to
solutes with molecular masses up to about 1500 Da [Hunter DR
et al (1976) J Biol Chem 251: 5069–5077]. Its occurrence is always
accompanied by depolarization, while onset of matrix swelling,
depletion of matrix pyridine nucleotides, outer membrane rupture
and release of intermembrane proteins including cytochrome c
depend on the open time. The PT is due to the reversible opening
of a high-conductance, voltage-dependent channel in the inner
mitochondrial membrane, the PT pore (PTP). In spite of many
efforts, its molecular identity remains unknown [reviewed in Ber-
nardi P et al (2006) FEBS J 273: 2077–2099]. In this lecture I

shall cover the essential aspects of PTP pathophysiology, with
specific emphasis on the role of matrix cyclophilin D [Giorgio V
et al (2010) Biochim Biophys Acta doi:10.1016/j.bbabio.2009.1 2.006];
Abstracts Invited speakers
6 FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies
the mechanism of action of cyclosporin A [Basso E et al (2008) J
Biol Chem 283: 26307–26311]; the modulation by the proton elec-
trochemical gradient [Bernardi P (1992) J Biol Chem 267: 8834–
8839] and redox effectors [Petronilli V et al (1994) J Biol Chem
269: 16638–16642]; and the consequences of PTP opening as a
key to understanding its role in cell dysfunction and death. From
this analysis the PTP emerges as a viable target for therapeutic
intervention in cancer [Rasola A et al (2010) FEBS Lett
doi:10.1016/j.febslet.2010.02.022] and degenerative diseases [Mer-
lini L et al (2008) Proc Natl Acad Sci USA 105: 5225–5229], and
a highly conserved event across species.
I13
Please view Plenary lectures
I14
Relationships between structure and functions
of a major complement inhibitor C4b-binding
protein
A. Blom
Laboratory Medicine Malmo
¨
, Section of Medical Protein
Chemistry, Lund Universty, Malmo
¨
, Sweden
The complement system is a vital component of innate immunity

and defends host from infections, alerts adaptive immunity and
clears the organism from unwanted debris such as dying cells,
misfolded proteins and immune complexes. Complement is
tightly regulated by a number of soluble and membrane bound
inhibitors and disturbances of this regulation are related to dis-
eases. C4b-binding protein (C4BP) is a major soluble complement
inhibitor composed of seven identical a-chains and a unique b-
chain, both of which contain complement control protein (CCP)
domains. All subunits are held together at their C-termini via
hydrophobic interactions between amphipathic helices and disul-
phide bridges. We have localized a number of binding sites for
C4BP ligands using recombinant mutants lacking domains, site-
directed mutagenesis, modelling and NMR. The binding site for
C4b, responsible for inhibition of complement, is localized to
CCP1-3 of a-chains and includes mainly positively charged aa.
An overlapping site is used by M proteins of Streptococcus pyog-
enes, an interaction that is one of the immune evasion mecha-
nisms of this pathogen. Overlapping binding sites are also used
by heparin and DNA. The C-terminus of a-chains including
CCP8 binds in turn some components of extracellular matrix and
amyloid. The high affinity (KD 0.2 nM) binding site for antico-
agulant protein S is localized to hydrophobic aa on CCP1 of the
b-chain. The C4BP-PS complex binds avidly to apoptotic cells
allowing their non-inflammatory phagocytosis.
I15
Transgenic plastids as expression factories in
biotechnology
R. Bock
Max-Planck-Institute of Molecular Plant Physiology, Potsdam,
Germany

Genetically modified plants (GM plants) can potentially provide
inexpensive production platforms for pharmaceuticals and nutra-
ceuticals. With the advent of technologies to alter the genetic
information inside plastids (chloroplasts), a new attractive target
for genetic engineering has become available to biotechnologists.
There are considerable attractions of the plastid genome as a
target for the expression of foreign genes. These include (i) the
plastids’ potential for high-level foreign protein expression
(Zhou et al., 2008; Oey et al., 2009), (ii) the possibility of trans-
gene stacking through expression of multiple genes from ope-
rons, and (iii) the absence of position effects and epigenetic gene
silencing mechanisms (Bock, 2007; Bock and Warzecha 2010).
From the biosafety perspective, the major attraction is the exclu-
sively or predominantly maternal inheritance of the plastid gen-
ome in most crop plants, greatly reducing the risk of
uncontrolled pollen spread of transgenes and thus allaying con-
cerns over environmental consequences of GM crop cultivation
(Ruf et al., 2007). Applications of chloroplast engineering in
basic research and biotechnology will depend critically on suc-
cess with extending the crop range of chloroplast transformation
(Ruf et al., 2001) and the feasibility to express transgenes in
non-green plastids (as present in fruits and tubers), which often
are less active in gene expression (Kahlau and Bock, 2008). The
state of the art in engineering the plastid genome of higher
plants will be described and selected applications in two areas of
biotechnology will be discussed: metabolic engineering and
molecular farming.
I16
Structure of chloroplast membrane
organization using cryo-electron tomography

E. Boekema, R. Kouril and G. Oostergetel
University of Groningen, Groningen, Netherlands
The thylakoid membrane architecture of chloroplasts was studied
by cryo-electron tomography (ET) and single particle electron
microscopy (EM). ET reconstructions of isolated, ice-embedded
specimens enabled to resolve the features of photosystem II
(PSII) in the native membrane of granal stacks and to get a close
view of its distribution. 3D analysis of subvolumes containing
PSII complexes provided a 3D structure of the PSII core complex
at 40 A
˚
resolution. Comparison with a recently proposed pseudo-
atomic model of the PSII supercomplex (EMBO Journal 28:
3052) revealed the presence of unknown protein densities right
on top of the four peripheral LHCII trimers. The positions of
individual PSII complexes were used to fit an entire membrane
layer with C2S2M2 supercomplexes. Fitting shows that many
supercomplexes must be of smaller size than C2S2M2 super-
complexes, to avoid overlap. The characteristic features of
PSII enabled assignment of the absolute orientation of individual
membranes of granal thylakoid discs and to get a detailed inter-
pretation of the membrane folding pattern. This shows that two
concentrically folded membranes make a unit of four layers. In
that unit, the outer membrane of the grana disc is interrupted on
only one side, whereas the inner membrane is almost closed. It will
be discussed how this is in line or in contrast with curent models.
I17
The genetic landscape of a cell
C. Boone
University of Toronto, Toronto, ON, Canada

A genome-scale genetic interaction map was constructed by
examining 5.4 million gene-gene pairs for synthetic genetic inter-
actions, generating quantitative genetic interaction profiles for
most genes in the budding yeast, Saccharomyces cerevisiae. A
network based on quantitative genetic interaction profiles reveals
a functional map of the cell in which genes of similar biological
processes cluster together in coherent subsets and highly corre-
lated profiles delineate specific pathways to define gene function.
The global network identifies functional cross connections
between all bioprocesses, mapping a cellular wiring diagram of
pleiotropy. Genetic interaction degree correlated with a number
of different gene attributes, which may be informative about
Invited speakers Abstracts
FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies 7
genetic network hubs in other organisms. An unbiased mapping
of the genetic landscape also provides a key for interpretation of
chemical-genetic interactions and drug target identification.
Finally, the identification of conditional essential genes, which
are essential within a specific genetic background, provides a
strategy for mapping genetic networks based upon natural varia-
tion that distinguishes two individuals.
I18
Genomics-enabled approaches for revealing
the molecular secrets of marine diatoms
C. Bowler
Ecole Normale Superieure, Plant Biology, Paris, France
Diatoms are eukaryotic photosynthetic microorganisms found
throughout marine and freshwater ecosystems that are responsi-
ble for around 20% of global primary productivity. A defining
feature of diatoms is their ornately patterned silicified cell wall,

which display species-specific nanoscale structures. These organ-
isms therefore play major roles in global carbon and silicon bio-
geochemical cycles. The two available whole genome sequences,
from Thalassiosira pseudonana and Phaeodactylum tricornutum,
provide a basis for comparative and functional genomics studies
of diatoms with other eukaryotes, as well as a foundation for
interpreting the ecological success of these organisms in a geno-
mic context. Of particular significance is the unprecedented pres-
ence of hundreds of genes from bacteria within diatom genomes.
The ancient origins of these gene transfers are testified by the
finding that more than 300 are found in both diatoms, and many
are likely to provide novel possibilities for metabolite manage-
ment and for perception of environmental signals. The wide
range of genetic resources that have been developed for P. tricor-
nutum (e.g. genetic transformation, RNAi, 130 000 ESTs) enable
the functions of these and other genes to be assessed using mod-
ern molecular methods. As a case in point I will show how gen-
ome-enabled resources can reveal how genes from different
origins have been recruited to ensure diatom survival in chroni-
cally iron-limited regions of the ocean.
I19
Structural and functional insights into
mitochondrial complex I
U. Brandt
Johann Wolfgang Goethe-University, Medical School, Frankfurt
am Main, Germany
Mitochondrial complex I is a membrane integral multiprotein
complex composed of 40+ different subunits and a total mass
of nearly 1 MDa. As entry point from NADH into the respira-
tory chain it serves a central function in energy metabolism and

couples electron transfer to ubiquinone with vectorial proton
translocation across the inner mitochondrial membrane. The
strictly aerobic yeast Yarrowia lipolytica is a powerful genetic
system to study complex I. By extensive structure based site-
directed mutagenesis we have analyzed the ubiquinone reducing
catalytic core of complex I that resides at the interface between
the 49-kDa and the PSST subunit of the peripheral arm of com-
plex I. In this functionally critical domain the ubiquinone is
reduced by cluster N
2
, the last in a chain of seven iron-sulfur
clusters. Our results define a likely entry pathway for ubiqui-
none leading to a region next to cluster N
2
, where the substrate
seems to bind to a fully conserved tyrosine. We could identify
residues responsible for the interaction of the isoprenoid side
chain of ubiquinone with complex I. Monitoring inhibitor sensi-
tivity allowed identification of binding regions for different clas-
ses of hydrophobic complex I inhibitors. New evidence on the
location of the ubiquinone binding pocket within complex I and
the path leading to the site where the hydrophobic substrate gets
reduced is presented. The molecular mechanism how complex I
couples proton translocation to electron transfer is still
unknown. We propose that long range conformational changes
drive proton pumping through a two-state stabilization change
mechanism involving distinct binding modes of charged ubiqui-
none intermediates.
I20
Molecular mechanism of photo-adaptation and

light entrainment of the circadian clock of
Neurospora crassa
M. Brunner
1
, E. Malzahn
1
, S. Ciprianidis
1
, T. Schafmeier
1
and
K. Kaldi
2
1
Biochemistry Center, Heidelberg University, Heidelberg,
Germany,
2
Physiology, Semmelweis University, Budapest, Hungary
The transcription factor White Collar Complex (WCC) is a core
element of the circadian clock of Neurospora crassa. White Col-
lar-1, a subunit of the WCC, is a blue-light photoreceptor
required for entrainment of the clock to photoperiods. Activated
WCC supports expression of Vivid (VVD), which is a negative
regulator of the WCC. WCC and VVD are both required for
photoadaptation of Neurospora. We present a molecular mecha-
nism how the photocycles of the flavin-binding LOV domains of
WC-1 and VVD allow photoadaptation to light intensities over
several orders of magnitude. We show that VVD is required for
the discrimination between day and night in naturally ambiguous
photoperiods with moonlight.

I21
Tuning the proton pumping stoichiometry in
cytochrome c oxidase by single mutations
P. Brzezinski
Stockholm University, Stockholm, Sweden
Cytochrome c oxidase (CytcO) is a multisubunit membrane-
bound enzyme, which catalyzes the oxidation of cytochrome
c and reduction of molecular oxygen to water. Part of the free
energy available from this reaction is used to pump one proton
across the membrane per electron transferred to oxygen. This
pumping stoichiometry of 1 H
+
/e
-
can be modulated by intro-
duction of single-site mutations, as far away as ~25 A
˚
from the
catalytic site, to obtain essentially any stoichiometry in the range
0–1. In many cases these mutations leave the intramolecular elec-
tron and proton-transfer rates unaffected. Understanding the
molecular mechanism underlying the partial or full uncoupling of
proton pumping from oxygen reduction is important for under-
standing the molecular design of the proton-pumping machinery
in the respiratory oxidases.
One key feature of all ‘uncoupled mutants’ studied to date is that
the pK
a
of an internal proton donor, Glu286, located deeply
within a proton pathway used for transfer of pumped and sub-

strate protons, is either increased or decreased (from 9.4 in the
wild-type oxidase). Because the Glu residue is found near the cat-
alytic site as well as near a putative exit pathway for pumped
protons, the altered pK
a
presumably reflects changes in the local
structural environment of the residue, which are important for
controlling the rates and trajectories of proton transfer to these
sites. These results establish a link between specific structural
changes in the proton pump and, its thermodynamic and kinetic
properties thereby offering mechanistic insights into the molecu-
lar design of the proton-pumping machinery of the oxidases.
Abstracts Invited speakers
8 FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies
I22
High resolution quantitative mass
spectrometry for analysis of proteomes and
PTMs
C. Choudhary
Department of Proteomics, University of Copenhagen, The NNF
Center for Protein Research, Copenhagen, Denmark
Proteins are fundamental constituents of all living organisms.
Mammalian genomes contain thousands of protein coding genes
(20 000 in human), and by alternative splicing of the mRNA,
and variance in gene sequences, they may encode many more
protein products. A great majority of proteins in eukaryotic cells
are modified by different posttranslational modification (PTMs).
PTMs are key constituents of cell signaling networks, compris-
ing some of the most sophisticated control switches in the cell.
Cells use various regulatory mechanisms such as dynamic regu-

lation of protein expression, PTMs, and protein-protein interac-
tions, to orchestrate cellular processes and to adapt to
environmental clues. Given the vast complexity and their all
essential role in controlling biology of living cells, analysis of
proteins and PTMs is one of the most exiting areas of research
in biology.
Recent developments in high-resolution mass spectrometry
(MS), computational proteomics, and optimized PTM-enrich-
ment strategies have greatly increased the depth of proteome
and PTM coverage by MS. Global phosphorylation analysis in
mammalian cells can routinely identify more than 10 000 phos-
phorylation sites in a single experiment. Large-scale analysis of
other PTMs such as lysine acetylation, ubiquitylation, and meth-
ylation are also becoming amenable to this technique. Quantita-
tive MS screens can provide unbiased, systems-wide information
about changes in protein expression, protein modifications, and
protein-protein interactions. I will discuss recent developments
in the quantitative MS, and highlight their pioneering applica-
tions in quantification of proteomes and posttranslational modi-
fications.
I23
The interplay between phosphorylation and
ubiquitination in regulating the innate immune
system
P. Cohen
University of Dundee, Medical Research Council Protein
Phosphorylation Unit, Dundee, UK
Infection by bacteria and viruses triggers cells of the innate
immune system to produce pro-inflammatory cytokines and inter-
ferons that mount the responses to fight the invading pathogens.

However these defence mechanisms are a double-edged sword
because the uncontrolled production of these substances causes
chronic inflammatory and autoimmune diseases. This talk will
focus on the signaling pathways downstream of Toll-Interleukin
Receptors. The engagement of these receptors by their agonists
triggers the formation of Lys63-linked polyubiquitin chains and
polyubiquitylated proteins, which then recruit and activate the
key protein kinases that drive the production of pro-inflamma-
tory cytokines and type 1 interferons. The mechanisms that initi-
ate the formation of Lys63-linked polyubiquitin chains and the
proteins that interact with them (NEMO, OPTN, ABIN1 and
ABIN2) will be discussed. Finally, the characterization of mice
that express an ABIN1 mutant unable to bind to Lys63-linked
polyubiquitin chains will be described and a model that accounts
for the striking phenotype displayed by these knock-in mice will
be presented.
I24
Functional interaction between chromatin
remodelers and non-coding RNA’s
D. Corona
Dipartimento di Biologia Cellulare e dello Sviluppo, Universita’
degli Studi di Palermo & Dulbecco Telethon Institute,
Palermo, Italy
The Hsr-x gene is developmentally expressed in almost all cells
types of D. melanogaster and is one of the most strongly induced
heat shock genes in flies. The Hsr-x locus encodes multiple non-
coding RNAs (ncRNA); the large nuclear species (Hsr-x-n) is
essential for the assembly and organization of hnRNP-containing
omega speckles. These special nuclear compartments are thought
to play essential roles in the storage/sequestration of members of

hnRNP family and other proteins playing important roles in
RNA maturation. ISWI is an evolutionarily conserved ATP-
dependent chromatin remodeler playing essential roles in chro-
mosome condensation, gene expression and DNA replication and
is also involved in a variety of nuclear functions including telo-
mere silencing, stem cell self-renewal, neural morphogenesis and
epigenetic reprogramming. Using an in vivo assay to identify fac-
tors that antagonize ISWI activity, we recovered a genetic inter-
action between ISWI and Hsr-x. Loss of Hsr-x function results
in a strong suppression of eye morphology and chromosome con-
densation defects caused by loss of ISWI activity. Moreover, the
organization of the omega speckles in ISWI mutant cells is pro-
foundly altered when compared to wild type cells. Interestingly,
immuno-FRISH analysis revealed a significant number of sites in
the nucleus where the chromatin-bound ISWI protein overlaps
the omega speckles-associated Hsr-x ncRNA. Remarkably,
RNAimmunoprecipitation assays, conducted in salivary glands
extracts, revealed a physical interaction between ISWI and Hsr-x
ncRNA.
I25
The bacterium Listeria monocytogenes: a
brilliant cell biologist and an insidious
biochemist
P. Cossart
Unite
´
des Interactions Bacte
´
ries-Cellules, Institut Pasteur, Paris,
France

Listeria monocytogenes is an ubiquitous bacterium that can con-
taminate food products and induce several forms of a disease
called listeriosis, an infection which is mortal in 30% of the
cases. Infection is mainly due to the capacity of the organism to
live and replicate in both phagocytic and non phagocytic cells. It
is also due to the property of the organism to be able to cross
three host barriers during infection, the intestinal barrier, the
blood brain barrier and in pregnant women the materno-fetal
barrier. A series of approaches have highlighted that this bacte-
rium has evolved amazing mechanisms to switch from saprohytic
life to virulence, to escape early innate host immune defense, to
invade non phagocytic cells, to spread from cell to cell and to
cross host barriers. We will focus the talk on recent results that
have highlighted new concepts in endocytosis/phagocytosis and
revealed.
Invited speakers Abstracts
FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies 9
I26
Molecular characterization of estrogen
receptor signaling in breast cancer cells
C. Zhao
1
, H. Gao
1
, N. Gustafsson
1
, Z. Papoutsi
1
, S. Jaffrey
1

,
J-A
˚
. Gustafsson
2
, J-A
˚
. Gustafsson
1
and K. Dahlman-Wright
1
1
Department of Biosciences and Nutrition, Karolinska Institutet,
Huddinge, Sweden,
2
Department of Biology and Biochemistry,
University of Houston, Houston, TX, USA
Breast cancer is a common malignancy worldwide. Although the
5-yr survival rate has increased with advances in detection and
treatment, many breast cancer patients still die from metastatic
disease. Therefore, more effective methods for diagnosis, classifi-
cation, treatment and individualized treatment are greatly
needed. Estrogen, acting via estrogen receptors (ERs), ERa and
ERb, has been well documented to play a critical role in the eti-
ology and progression of breast cancer. The introduction of
anti-estrogens such as tamoxifen for the treatment of breast can-
cer constitutes a milestone in the treatment of this disease. How-
ever, many tumors are not responsive to this treatment or
become resistant over the cause of the treatment. Our research is
focused on understanding aspects of estrogen signaling at the

molecular level including how estrogen signaling can be modu-
lated by other pathways than ligand-binding. To increase our
understanding of the molecular mechanism of estrogen signaling
in breast cancer cells, we determined the global DNA-binding
profile ERa and ERb and showed that there were differences in
the DNA-binding profiles between these two receptors including
differences with regard evolutionary conservation, distance to
transcription start sites and binding site characteristics. We have
shown that ERa signaling and proliferation can be inhibited by
the ERb variant ERb2 through proteasome-dependent degrada-
tion of ERa and enhanced by the RBCK1 protein, the latter
might involve direct regulation of the ERa promoter. More
recently we have focused on crosstalk between AP-1 and ERb
signaling showing that inhibition of AP-1 signaling decreases
ERb DNA-binding and signaling.
I27
On the role of protons and O
2
partial pressure
in water oxidation by photosystem II
H. Dau, I. Zaharieva, M. Haumann, J. Wichmann, A. Grundmeier,
A. Klauss and L. Gerencser
Freie University Berlin, FB Physik, Berlin, Germany
In plants and cyanobacteria, two water molecules are split into
four energized electrons (reducing equivalents), four protons and
one dioxygen molecule. This process is driven by light and facili-
tated by a manganese-calcium complex bound to protein subunits
of photosystem II (PSII), the latter being an impressive cofactor-
protein complex embedded in the thylakoid membrane separating
the lumen and stroma compartment. Experimental results on the

‘backpressure’ by the reaction products ‘proton’ (low pH at the
lumen side of PSII) and ‘oxygen’ (high O
2
partial pressure) are
presented. The results relate to the mechanism of water oxidation
in PSII, specifically to the spatio-temporal coordination of elec-
tron and proton transfer at the donor side of PSII. Implications
for the function of PSII in intact organisms also will be discussed.
I28
Exocytosis, endocytosis and lysosomal
targeting of the Aquaporin-2 water channel: an
interplay of phosphorylation and
ubiquitination
P. Deen
Physiology, Radboud University Nijmegen Medical Center,
Nijmegen, Netherlands
Regulation of a proper water homeostasis is essential for terres-
trial animals to survive. This regulation is mainly achieved by
drinking and modulation of reabsorption of water from pro-urine,
of which the latter is under control of the anti-diuretic hormone
vasopressin. In states of hypernatremia or hypovolemia, vasopres-
sin is released from the pituitary, binds its type-2 receptor in the
kidney collecting duct and induces the apical surface expression of
aquaporin-2 water channels upon activation of a cAMP signaling
cascade. When AVP levels drop, but also with additional vaso-
pressin-counteracting hormones, AQP2 is internalized for recy-
cling or degradation. Using polarized cells or renal material, we
found that forskolin/vasopressin increases Ser256 phosphoryla-
tion (pS256) of AQP2, which is essential for its translocation to
the apical membrane. This translocation was reversed upon

removal of forskolin or co-incubation with vasopressin-counter-
acting hormones, which did not involve pS256 dephosphorylation,
but essentially involved a transient increase in AQP2 K63-linked
ubiquitination with 2–3 ubiquitin moieties on Lys270 only in
AQP2. This ubiquitination was an essential process, as AQP2-
K270R, which cannot be ubiquitinated, was impaired in its endo-
cytosis. AQP2 stably fused to ubiquitin (AQP2-Ub) localized to
lysosomes. Ubiquitin-dependent endocytosis of AQP2 was fol-
lowed by AQP2 phosphorylation at S261, which seems to stabilize
AQP2 ubiquitination and endocytosis. Following endocytosis,
AQP2 is targeted for degradation to multivesicular bodies and
lysosomes, which involves interaction with the MIT1 domain of
LIP5, which is part of the ESCRTIII complex.
I29
Ubiquitin-mediated regulation of NF-jB
activation and autophagy
I. Dikic
Institute of Biochemistry II, Goethe University Frankfurt, Germany
An increasing number of distinct functions have been assigned to
different types of ubiquitin modifications (mono ubiquitin versus
ubiquitin chains). In these processes Ub acts as a signalling com-
ponent able to trigger molecular events in cells. Structural studies
have revealed significant differences between ubiquitin chains of
Lys48, Lys63 and linear linkages. We have recently shown that
the UBAN domain of NF-jB essential modulator (NEMO) binds
specifically to linear Ub chains and that this step is essential for
NF-jB activation. These findings explain the detrimental effect of
NEMO mutations in patients suffering from X-linked ectodermal
dysplasia and immunodeficiency. More recently, we have identi-
fied novel components of the linear ubiquitin-dependent pathways

that control the NF-jB and apoptotic pathways downstream of
TNF receptors. Removal of harmful protein aggregates, damaged
organelles and microbes is mediated by autophagy, a process by
which the cell sequesters cytosolic cargo and delivers it for degra-
dation by the lysosome. The involvement of ubiquitin in selective
autophagy is becoming more evident: autophagic clearance of
protein aggregates requires specific autophagy receptors. Identifi-
cation of p62/SQSTM1 and NBR1 as autophagy receptors, which
simultaneously bind both ubiquitin and autophagy-specific ubiqu-
itin-like modifiers, LC3/GABARAP, has provided a molecular
Abstracts Invited speakers
10 FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies
link between ubiquitination and autophagy. The molecular details
of selective autophagy suitable for clearance of various cargoes,
ranging from ubiquitinated protein aggregates to membrane-
bound organelles and microbes will be discussed.
I30
Abstract missing, please view Abstract Addendum
I31
The Structural basis for ion conduction and
gating in pentameric ligand-gated ion
channels
R. Dutzler
Department of Biochemistry, University of Zurich, Zurich,
Switzerland
The pentameric ligand-gated ion channels (pLGICs) constitute a
family of neurotransmitter receptors that are key players in the
control of electric signaling at chemical synapses. The family
codes for a conserved scaffold of channel proteins that open in
response to the binding of neurotransmitter molecules. We have

determined the X-ray structures of two prokaryotic family mem-
bers from the bacterium Erwinia chrysanthemi (ELIC) at 3.3 nm
resolution [1] and from the bacterium Gloeobacter violaceus
(GLIC) at 3.1 nm resolution [2]. Both proteins form cation selec-
tive channels and bear most of the structural hallmarks of the
family including the N-terminal extracellular ligand-binding
domain and the four helices of the pore domain. Despite the
overall similarity, both structures adopt distinct conformations of
the ion conduction pathway. The structure of ELIC shows a
non-conductive state with rings of hydrophobic residues at the
extracellular side of the pore preventing ion permeation. This
hydrophobic barrier has opened in the structure of GLIC to a
funnel shaped pore, where a ring of conserved glutamate residues
at the intracellular constriction creates an ion-coordination site.
GLIC is thus believed to represent a conducting conformation of
the channel. In combination, both structures suggest a novel gat-
ing mechanism for pentameric ligand-gated ion channels where
channel opening proceeds by a change in the tilt of the pore-
forming helices. The structures thus provide a first detailed view
into how a pLGIC may open and selectively conduct ions.
References:
1. Hilf RJC, Dutzler R. Nature 2008; 452: 375–379.
2. Hilf RJC, Dutzler R. Nature 2009; 457: 115–118.
I32
Lessens from the pH-sensitivity of the rate of
peptidyl transfer from P-site to A-site tRNA on
the translating ribosome
M. Ehrenberg
1
, M. Johansson

1
, I. Kaweng
1
, P. Strazwski
2
,
M. Pavlov
1
and J. Aqvist
1
1
Uppsala University, Dept of Cell and Molecular Biology, Uppsala,
Sweden,
2
Universite
´
Claude Bernard, Lyon 1, Institut de Chimie et
Biochimie Mole
´
culaires, Lyon, France
We studied the pH-dependence of ribosome catalyzed peptidyl
transfer from fMet-tRNAfMet to the aa-tRNAs Phe-tRNAPhe,
Ala-tRNAAla, Gly-tRNAGly, Pro-tRNAPro, Asn-tRNAAsn
and Ile-tRNAIle, selected to cover a large range of intrinsic pKa-
values for the a -amino group of their amino acids. The peptidyl
transfer rates were different at pH 7.5 and had different apparent
pKa-values, defined as the pH-value at which the rate was half
maximal. The apparent pKa-values were downshifted in relation
to those of the aa-tRNAs in solution. Gly-tRNAGly had the
smallest (about 0.25 pH unit), while Ile-tRNAIle and Ala-

tRNAAla had the largest (about 1.5 pH unit) downshifts. These
down shifts correlate strongly with molecular dynamics (MD)
estimates of the down shifts in pKa-values of the alpha-amino
groups of these aa-tRNAs upon A-site binding. Our data demon-
strate the chemistry of peptide bond formation, rather than
accommodation, to be rate limiting for peptidyl transfer at pH
7.5 in the Gly and Pro cases and strongly suggest rate limiting
chemistry for all aa-tRNAs.
I33
Lipidomics in health & disease
K. Ekroos
Zora Biosciences, Espoo, Finland
Lipidomics is defined as the systems-level analysis of lipid species,
their abundance, biological activity, as well as sub-cellular locali-
zation and tissue distribution. Technological advances in mass
spectrometry and associated method development have produced
lipidomic techniques capable in characterizing and quantifying
hundreds of molecular lipid species directly in total lipid extracts.
Moreover, the high detection precision at the molecular lipid
level prevents false-positive identifications, a necessity for quality
lipid research and enables studies requiring GLP. The identifica-
tion of lipid metabolic dysfunctions responsible for disease
pathology also requires the coupling of customized bioinformatic
tools to the analytical chemistry of lipidomics. Molecular lipids
are the window to meaningful biological insight and a prerequi-
site for piecing together the molecular mechanisms of health and
disease processes. Finally, molecular lipidomics is a valuable
addition to the toolkits used both in biomarker discovery and in
the identification of new drug targets.
I34

Probing intracellular kinetics at the level of
single molecules
B. English, A. Sanamrad, H. Vasili and J. Elf
Uppsala university, Uppsala, Sweden
I will present our resent advancements in tracking individual
freely diffusing fluorescent protein molecules at high time resolu-
tion in the cytoplasm of bacterial cells. In vivo tracking of indi-
vidual proteins molecules makes it possible to study kinetics high
time resolution without synchronizing the population of mole-
cules. For example by monitoring the kinetics of the response
mediator RelA we have developed a single molecule assay to
study stress response and starvation at the level of individual
bacteria. The RelA protein binds to a small fraction of ribo-
somes, where it synthesizes the global transcriptional regulator
ppGpp in response to amino acids deprivation. This the ppGpp
molecule binds to the RNAP and rapidly reprograms the cell for
the new environment, in what is called the stringent response.
While Escherichia coli contains on average about 100 RelA mole-
cules and 10 000 ribosomes, using a photo-activatable fluorescent
probe we can activate only a few fluorescent molecules per cell at
any given time and track them at high time resolution. When the
cell grows exponentially, RelA trajectories closely resemble trajec-
tories of fluorescently tagged ribosomal proteins (D~0.4 lm
2
/s as
compared to D~0.3 lm
2
/s for ribsomes). After nutritional down-
shift, RelA binding kinetics changes rapidly and the protein
diffuses very fast (D~3.5 lm

2
/s) as if it only binds to ribsomes
transiently. The assay has made it possible to study the rapid
and transient stringent response in individual cell as well as the
heterogeneity in the stress response over the population.
Invited speakers Abstracts
FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies 11
I35
Human brown adipose tissue
S. Enerba
¨
ck
University of Gothenburg, Go
¨
teborg, Sweden
Scarcity of food and cold weather have accompanied us and con-
stituted serious threats to our survival for the entire duration of
our species’ history. Fortunately, two tissues have been selected
by nature to help us deal with these difficulties: white adipose tis-
sue (WAT) and brown adipose tissue (BAT). While WAT helps
us compensate for fluctuating availability of energy, BAT keeps
us warm – at least during our first days of extra-uterine life. As
will be discussed, it is most likely of importance to us for longer
than that. It is only very recently that, by having an adequate
food supply and modern housing, we have circumvented the
problems associated with famine and cold weather – at least in
some parts of the world – and thus become less dependent on
WAT and BAT for our survival. This has occurred to the extent
that new dangers have been created. A sedentary inactive lifestyle
and a wealth of calories have generated a cluster of obesity-

related health problems known as the metabolic syndrome or
syndrome X. Which in turn is associated with a positive energy
balance; here BAT can play a role by increasing energy expendi-
ture. BAT is only present in mammals and is estimated to have
emerged some 150 million years ago. BAT is unique in the sense
that brown adipocytes can uncouple respiration from ATP syn-
thesis. A role for metabolically active BAT in the normal physiol-
ogy of healthy adults will be discussed as well as potential
therapeutic implications.
I36
Abstract missing, please view Abstract Addendum
I37
Mechanisms of initiation of DNA replication in
human mitochondria
M. Falkenberg
Institute of Biomedicine, University of Gothenburg, Gothenburg,
Sweden
Mitochondrial DNA is replicated by a unique enzymatic machin-
ery, which is distinct from the replication apparatus used for
copying the nuclear genome. In combination, DNA polymerase
gamma and TWINKLE DNA helicase form a processive replica-
tion machinery, which can use dsDNA as template to synthesize
single-stranded DNA molecules and addition of the mitochon-
drial ssDNA-binding protein stimulates the reaction further.
Recently, we have studied the mechanisms of origin-specific initi-
ation of lagging-strand DNA synthesis. We found that the mito-
chondrial RNA polymerase (POLRMT) is the primase required
for initiation of DNA synthesis from the light-strand origin of
DNA replication (OriL). Using only purified POLRMT and
DNA replication factors, we could faithfully reconstitute OriL-

dependent initiation in vitro. Leading-strand DNA synthesis is
initiated from the heavy-strand origin of DNA replication and
passes OriL. The single-stranded OriL is exposed and adopts a
stem-loop structure and at this stage, POLRMT initiates primer
synthesis from the single-stranded loop region. After about 25 nt,
POLRMT is replaced by DNA polymerase gamma, and DNA
synthesis commences. Our findings demonstrate that POLRMT
can function as an origin-specific primase in mammalian mito-
chondria.
In ongoing studies, we are trying to understand how mtDNA
replication is initiated at the leading-strand origin of mtDNA
replication, OriH. We will present data suggesting a new mecha-
nism for transcription termination and primer formation in
human mitochondria.
I38
Conformational changes seen in crystal
structures of oxidized, reduced and mutant
forms of cytochrome c oxidase: clues to
coupling and gating mechanism?
S. Ferguson-Miller, L. Qin, J. Liu, C. Hiser and F. Li
Michigan State University, Biochemistry and Molecular Biology,
East Lansing, MI, USA
There is increasing recognition that altered mitochondrial meta-
bolism and energy production is involved in many disease states,
particularly those associated with neurological dysfunction, aging
and obesity. Cytochrome c oxidase is a critical controlling
enzyme in the mitochondrion, but despite intensive study, impor-
tant aspects of its mechanism and regulation remain elusive. We
study a bacterial homolog of the mammalian mitochondrial
enzyme, which has a simpler structure, is readily mutated, and is

amenable to crystallographic analysis. Our recent high-resolution
crystal structures provide new insights into its ligand binding
behavior (Qin et al, Biochemistry 2008; 47: 9931) and new evi-
dence of redox-linked conformational change (Qin et al, Bio-
chemistry 2009; 48: 5121). An exciting aspect of these results is
that the changes seen upon reduction of the enzyme suggest a
conformationally-controlled gating mechanism that has not pre-
viously been considered. Another interesting finding is that lipid
and steroid binding sites are conserved from bacterial through to
mammalian oxidases, suggesting functional importance. Crystal
structures of mutant forms in proton uptake pathways show
minimal alteration in structure except for missing key water
molecules, but reveal similar redox-linked conformational
changes as wildtype. Spectral changes in crystals during x-ray
irradiation reveal evidence of a strained configuration in the fro-
zen state. The importance of conformational change and lipidic
ligands in the mechanism and regulation of cytochrome oxidase
is a focus of our current studies. (NIH GM26916; MSU REF 03-
016).
I39
Nature’s way to exploit dihydrogen as an
alternative fuel
B. Friedrich
Humboldt University Berlin, Institute of Biology/Microbiology,
Berlin, Germany
Hydrogen, an attractive microbial energy source, is released into
the anoxic environment by fermentation where it gets immedi-
ately consumed by anaerobic organisms. Both hydrogen evolu-
tion and hydrogen consumption are catalyzed by metal-cofactor
containing hydrogenases. In order to use these catalysts for effi-

cient production of sunlight-driven hydrogen from water, a num-
ber of requirements have to be met. Particular challenges are: (i)
tight coupling of the electron flow between the algal or cyanobac-
terial photosystems and the hydrogenase protein and (ii) toler-
ance of the hydrogenase towards oxygen that unavoidably
escapes during the water splitting process. The majority of hy-
drogenases are sensitive to oxygen and either irreversibly dam-
aged or inactivated by oxygen. A few [NiFe] hydrogenases,
resident in aerobic hydrogen-oxidizers, tolerate ambient oxygen
during catalysis. The molecular background of oxygen tolerance
is diverse and may depend on the access of oxygen to the
active site, on the interplay between various metal cofactors, their
Abstracts Invited speakers
12 FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies
protein environment and their redox potentials. Insights into
these mechanisms open new strategies for genetic engineering of
an optimal hydrogenase catalyst suitable for biotechnological
applications. Some account of this work has been published in a
recent review article. (O. Lenz, M. Ludwig, T. Schubert, I. Bu
¨
r-
stel, S. Ganskow, T. Goris, A. Schwarze, B. Friedrich. H
2
con-
version in the presence of O
2
as performed by the membrane-
bound [NiFe]-hydrogenase of Ralstonia eutropha. ChemPhy-
sChem (2010) DOI:10.1002/cphc.200901002.)
I40

Abstract missing, please view Abstract Addendum
I41
From hot to cool and more for less: new
developments for structural biology
E. Garman
Biochemistry, University of Oxford, Oxford, UK
Structural biology relies on X-ray crystallography to provide
much of the three dimensional information on macromolecules
that informs biological function. To enable problems not previ-
ously accessible to structure solution to be tackled, improved
methods must be developed. A notable example of this has been
the progress in finding protocols to cryocool protein crystals
prior to data 100K collection to reduce the rate of radiation
damage by around a factor of 70 compared to that at room tem-
perature (RT): from hot to cool and more for less. Radiation
damage to the sample is an inherent problem when utilising ion-
ising X-radiation in MX, and it is now known that radiation
damage can also be a limiting factor for MX at 100K. Following
our measurement of 30 MGy (1 Gy = 1 J/kg energy absorbed)
for the experimental dose limit for 100K protein crystals, we
tried to determine a limit for RT samples. The unexpected
results will be presented. Current ongoing methods investigations
that will be described include studies of 100K and RT radiation
damage in macromolecular crystals in order to inform both our
understanding and putative mitigation strategies, and trace ele-
mental analysis of liquid and crystalline proteins using micro-
PIXE (particle induced X-ray emission), allowing determination
of their stoichiometric ratio to an accuracy of between 10% and
20%.
I42

Transport through nuclear pore complexes
S. Frey
1
,B.Hu
¨
lsmann
1
, A. Labokha
1
, C. Ader
2
, M. Baldus
2
and
D. Go
¨
rlich
1
1
Max Planck Institute for Biophysical Chemistry, Cellular
Logistics, Go
¨
ttingen, Germany,
2
Bijvoet Center for Biomolecular
Research, Utrecht University, Utrecht, Netherlands
The permeability barrier of nuclear pore complexes (NPCs) is a
passive and yet highly efficient sorting device that controls all
exchange between nucleus and cytoplasm. It suppresses the flux
of inert macromolecules >30 kDa, but also allows rapid passage

of even very large cargoes, provided these are bound to appropri-
ate nuclear transport receptors (NTRs). FG-repeat domains bind
NTRs during facilitated NPC passage and constitute the crucial
elements of this barrier. They are essential for viability and com-
prise up to 50 repeat units. Each unit contains a hydrophobic
cluster, typically of the sequence FG, FxFG or GLFG, sur-
rounded by more hydrophilic spacer sequences. We observed that
FG-repeat domains form FG-hydrogels. These gels are fascinat-
ing materials that display permeability properties very similar to
those of authentic NPCs, allowing an up to 20,000-fold faster
entry of large NTRdcargo complexes as compared to the cargoes
alone. While supporting massive importin- or exportin-mediated
cargo influx, such gels remain firm barriers towards inert objects
that lack nuclear transport signals. This indicates that FG-hydro-
gels reseal immediately behind a translocating species and thus
possesses self-healing properties. The presentation will further
address the following questions: What is the molecular and struc-
tural basis of barrier formation? Why do the meshes of the bar-
rier open at least 100 times faster in the immediate vicinity of a
nuclear transport receptor than elsewhere in the gel? How do FG
repeat domains behave on a nanoscopic scale, i.e. in authentic
NPCs?
I43
Abstract missing, please view Abstract Addendum
I44
High-resolution imaging of neurotransmitter
receptors: learning from a single molecule
L. Groc
CNRS – Universite
´

Bordeaux, Bordeaux, France
In the nervous system, the trafficking of neurotransmitter receptor
in and out synapses has emerged as a key process through which
synapses adapt their strength in response to salient environmental
cues. Although numerous molecules and signalling cascades have
been identified over the last decades, our current understanding
of the molecular dynamics that sustain synaptic adaptations is
still rather limited. The membrane insertion and endocytosis of
neurotransmitter receptors mostly regulate the number of surface
receptor in various membrane compartments. Using single mole-
cule and particle tracking it recently emerged that surface neuro-
transmitter receptors are highly dynamic. Such observation
required the development of new experimental approaches to spe-
cifically label and track receptor with appropriate time- and
space-resolutions. Physiologically, it unravelled that the lateral
diffusion of glutamatergic receptors, which mediate most excit-
atory drive in the brain, plays a major role in regulating receptor
trafficking to and from synaptic sites as well as the short- and
long-term adaptations of excitatory synapses. These develop-
ments, i.e. how studying single surface receptor at the nano-scale
level provided fundamental knowledge about the synaptic and
neuronal plasticity, will be discussed in this presentation.
I45
Mechanistic insights into complement
activation and regulation
P. Gros
Crystal and Structural Chemistry, Utrecht University, Utrecht,
Netherlands
The complement system is a crucial part of the innate immunity
in mammals. This system is formed by ~30 plasma proteins and

cell-surface receptors, which enable the host to clear invading
pathogens and altered host cells, while protecting healthy host
tissue. Through structural studies we have revealed the molecular
mechanisms responsible for the amplification and regulation of
the central steps in the complement pathways. Structures of the
central complement component C3 (1641 res.) and its activated
form C3b revealed intricate domain arrangements and marked
conformational changes that lead to covalent labelling by C3b,
which mark target cells for immune clearance [1,2]. Activation is
performed by labile protease complexes, i.e. C3 convertases,
Invited speakers Abstracts
FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies 13
which are formed on the target cell surfaces. We determined
structures of the pro-enzyme factor B [3], the C3 pro-convertase
[4] and the active short-lived (half-life time ~90 s) C3 convertase
[5]. Furthermore, we determined structures of C3b in complex
with complement regulators that protect host cells from comple-
ment activation [6]. Together our data provide comprehensive
insights into the formation, activity, specificity and regulation of
the central C3 convertase, which cleaves C3 into the active C3b
and thereby amplifies the complement response in immune clear-
ance.
References:
1. Janssen et al. Nature 2005; 437: 505–511.
2. Janssen et al. Nature 2006; 444: 213–216.
3. Milder et al. Nature SMB 2007; 14: 224–228.
4. Janssen et al. EMBO J 2009; 28: 2469–2478.
5. Rooijakkers, Wu et al. Na Immun 2009; 10: 721–727.
6. Wu et al. Nat Immun 2009; 10: 728–733.
I46

Please view Plenary lectures
I47
Systematic analysis of dynamic signaling
modules by quantitative mass spectrometry
M. Gstaiger, M. Varjosalo, A. Wepf, T. Glatter, O. Rinner,
R. Aebersold and A. van Drogen
ETH, IMSB, ETH Ho
¨
nggerberg, Zu
¨
rich, Switzerland
Biological processes such as metabolism and signal transduction
emerge from dynamic networks of interacting protein complexes.
Affinity purification combined with mass spectrometry (AP-MS)
represents the most widely used approach for studying protein
complexes under near physiological conditions. However compre-
hensive analysis of protein complexes that constitute entire sig-
naling systems in human cells is limited by a number of
experimental challenges. These limitations are primarily linked to
the lack of time resolved quantitative MS data on transient sig-
nalling dependent protein-protein interactions, problems in iden-
tifying the true protein complex composition from AP-MS data
and the low throughput of existing AP-MS workflows. We have
introduced a combined computational and mass spectrometric
framework for the quantitative analysis of changes in protein
complex composition and protein modifications. We will also
report on a recently developed strategy for the systematic analy-
sis of human interaction proteomes at increased through put.
When combined these approaches mark a significant step for-
ward towards deciphering dynamic networks of human signalling

complexes. Recent applications of these methods for the analysis
of cellular signaling modules that control of cell growth will be
discussed.
I48
Abstract missing, please view Abstract Addendum
I49
The dynamic amyloid landscape
P. Hammarstrom
IFM-Department of Chemistry, Linkoping University, Linkoping,
Sweden
We investigate conformational heterogeneity and templated con-
formational conversion of amyloidogenic proteins associated with
human diseases. We study the human prion protein, the amyloid
beta peptide and transthyretin which are all linked to protein
deposition in the form of amyloid in vivo. There is ample evi-
dence that different point mutations in these proteins dictate the
disease phenotype. For the human prion protein around 20 dif-
ferent point mutations and two SNPs (rendering single residue
substitutions) have been described which all present different dis-
ease phenotypes. The same phenomenon holds true for transthy-
retin (>100 mutations have been identified) and for amyloid
beta which also extensively varies in peptide chain length,
depending on processing. Our goal is to understand the prerequi-
sites for amyloid fibril formation and conformational stability of
both the native structure and the misfolded conformations. We
have found that several self-assembled forms of these proteins
can interconvert emphasizing that these are dynamic structures
with rather shallow barriers between them.
References:
1. Almstedt K et al. Amyloid fibrils of human prion protein are spun

and woven from morphologically disordered aggregates. Prion
2009; 3(4): 224–235.
2. So
¨
rgjerd K. et al. Prefibrillar transthyretin oligomers and cold
stored native tetrameric transthyretin are cytotoxic in cell culture.
Biochem Biophys Res Commun 2008; 377(4):1072–1078.
3. Nilsson KP et al. Imaging distinct conformational states of amy-
loid-beta fibrils in Alzheimer’s disease using novel luminescent
probes. ACS Chem Biol 2007; 2(8):553–560.
I50
Abstract missing, please view Abstract Addendum
I51
Extensive crosstalk between O-GlcNAcylation
and phosphorylation: a new paradigm for
cellular signaling?
G. Hart
Biological Chemistry, Johns Hopkins University School of Medi-
cine, Baltimore, MD, USA
The cycling of O-linked b-D-N-acetylglucosamine (O-GlcNAc)
at serine or threonine moieties is abundant on nuclear and cyto-
plasmic proteins, where it serves as a nutrient/stress sensor to
regulate signaling, transcription and cellular metabolism. Several
studies have shown that a modest increase in global O-GlcNAcy-
lation affects phosphosite occupancy at nearly every actively
cycling phosphorylation site. Recently, in collaboration with
Don Hunt’s group at the University of Virginia, we developed a
chemico-enzymatic photochemical enrichment method, combined
with electron transfer dissociation (ETD) mass spectrometry, that
allows, for the first time, detection of O-GlcNAc site occupancy

on nucleocytplasmic proteins at a level of sensitivity comparable
to that possible for phosphorylation. Using this method to study
signaling processes that regulate cell division, we found that the
crosstalk between site specific phosphorylation and O-GlcNAcy-
lation is surprisingly extensive, with nearly every actively cycling
phosphorylation site affected by altered O-GlcNAcylation. Major
known signaling cascades that regulate cell division are strikingly
affected by a modest change in O-GlcNAcylation. Our studies
suggest that O-GlcNAcylation generally modulates many signal-
ing cascades and phosphate-mediated molecular switches to
allow them to be highly responsive to nutrients and stress. Sup-
ported by NIH R01 CA42486 and DK61671. Dr. Hart receives a
share of royalty received by the university on sales of the CTD
110.6 antibody. Terms of this arrangement are managed by
JHUSOM.
I52
Abstract missing, please view Abstract Addendum
Abstracts Invited speakers
14 FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies
I53
Ligand recognition and plasticity in HIV
envelope glycoprotein gp120
W. A. Hendrickson
1
, H. Xie
2
, M. Le-Khac
2
, A. Korkut
3

,D.Ng
4
,
J. R. Courter
4
, Y. D. Kwon
5
, A. Scho
¨
n
6
, N. Madani
7
,J.M.
LaLonde
8
, I. M. Chaiken
9
, E. Freire
6
, J. Sodroski
7
,P.D.
Kwong
5
and A. B. Smith
4
1
Department of Biochemistry and Molecular Biophysics, Columbia
University, Howard Hughes Medical Institute, New York, NY,

USA,
2
Department of Pharmacology, Columbia University, New
York, NY, USA,
3
Department of Biochemistry and Molecular Bio-
physics, Columbia University, New York, NY, USA,
4
Department
of Chemistry, University of Pennsylvania, Philadelphia, PA, USA,
5
National Institutes of Health, Vaccine Research Center, Bethesda,
MD, USA,
6
Department of Biology, Johns Hopkins University,
Baltimore, MD, USA,
7
Department of Cancer Immunology and
AIDS, Dana-Farber Cancer Institute, Boston, MA, USA,
8
Chemis-
try Department, Bryn Mawr College, Bryn Mawr, PA, USA,
9
Department of Biochemistry and Molecular Biology, Drexel
University College of Medicine, Philadelphia, PA, USA
Crystallographic analyses of core portions of HIV gp120 glyco-
proteins showed how these viral envelope proteins bind to human
CD4, the primary cellular receptor for HIV, and to a neutralizing
antibody fragment, Fab 17b, whose binding epitope overlaps
with that of HIV co-receptors CCR5 and CXCR4. HIV gp120

has a substantial cavity next to CD4 residue Phe43 at the focus
of protein-protein interaction. Thermodynamic analyses indicated
that gp120 undergoes remarkable conformational ordering when
it binds CD4 or 17b. We subsequently performed computational
analyses of the intrinsic flexibility of core gp120. We also
designed the F43C mutant variant of the D1D2 portion of CD4
for addition of chemical moieties in place of the Phe43 side
chain, and we analyzed structure-activity relations from the bind-
ing of a library of thiol-reactive compounds that produced
D1D2-F43C-X derivatives. HIV gp120 bound more tightly to
some of these derivatives than to wild-type CD4. We determined
crystal structures for five derivatized D1D2 proteins in complexes
with core gp120 and Fab 17b. Chemical entities bound into the
Phe43 cavity, which expanded in response to ligands. This plas-
ticity in gp120 was associated with modified binding properties in
cellular assays. We have also studied properties of compounds
derived from the gp120-ligand NBD-556, showing thermody-
namic characteristics similar to CD4 binding. Crystals prepared
from a core gp120 complexed with Fab 48d, but without CD4,
can bind these compounds. Crystal structures of several such
complexes reveal binding into the Phe43 cavity as for the D1D2-
F43C-X derivatives.
I54
What mitochondria learned from their
bacterial ancestors: oxidation-driven protein
folding
J. Herrmann
University of Kaiserslautern, Cell Biology, Kaiserslautern,
Germany
In the bacterial periplasm and in the ER of eukaryotic cells, sulf-

hydryl oxidases catalyze the formation of disulfide bridges
between cysteine residues in order to induce or stabilize protein
folding. In contrast, other cellular compartments are assumed to
be generally counteracting the formation of disulfide bridges to
maintain proteins in a reduced state. It therefore was completely
unexpected when recently a machinery was identified in the inter-
membrane space of mitochondria that catalyzes the oxidative
folding of proteins. This machinery is essential for protein trans-
location of certain cysteine-containing precursor proteins and
consists of two components which are highly conserved and
essential for viability: The oxidoreductase Mia40 and the sulfhy-
dryl oxidase Erv1. Mia40 serves as import receptor that cova-
lently binds to proteins after their passage through the TOM
complex, thereby converting them to their oxidized state. Since
only unfolded, reduced proteins can traverse the TOM channel,
this irreversibly traps the imported proteins in the mitochondria.
Finally, Mia40 is reoxidized by Erv1. Thus, the principle to oxi-
dize proteins was presumably conserved from the bacterial peri-
plasm to the intermembrane space of mitochondria, but this
system was adapted during evolution so that it now mediates the
vectorial translocation of proteins from the cytosol into the orga-
nelle.
I55
Teaching old enzymes new tricks
D. Hilvert
Laboratorium fu
¨
r Organische Chemie, ETH Zu
¨
rich, Zu

¨
rich,
Switzerland
While nature evolves its catalysts over millions of years, enzyme
engineers try to do it a bit faster. Enzyme active sites provide
highly optimized micro-environments for the catalysis of biologi-
cally useful chemical transformations. Consequently, changes at
these centers can have large effects on enzyme activity. The pre-
diction and control of these effects provides a promising way to
access new functions. In this talk, several strategies for tailoring
the properties of existing enzymes will be presented, together with
the insights into function and evolution that emerge from such
studies.
I56
Progress in production and purification of
proteins
S. Hober
Biotechnology/KTH, Proteomics, Stockholm, Sweden
The demand for high-throughput recombinant protein produc-
tion has increased markedly with the increased activity in the
field of proteomics. Depending on the aim of the produced target
protein different measures are needed. For high throughput pro-
duction and purification of many proteins in parallel, general
methods without the need for optimization for each target pro-
tein are desirable. Here, strategies for optimization of protein
production as well as methods for analysis of protein levels in
cell cultures will be discussed. Moreover different strategies for
improvement of selectivity and robustness of purification strate-
gies will be presented both for protein affinity chromatography
and ion-exchange chromatography. Novel handles for improved

selectivity of ion exchange protein purification processes have
been developed and successfully used as genetically fused purifi-
cation handles both in anion and cation exchange protein purifi-
cation. A recently created purification tag with dual affinity will
be presented. This tag allows for orthogonal affinity purification.
High-throughput protein production and analysis methods for
parallel production and MS analysis in 96-plate format will be
presented. Moreover high throughput methods to render purifica-
tion of hundreds of proteins weekly possible have been developed
as well as strategies for solid phase refolding and proteolytic
removal of the purification domains.
Invited speakers Abstracts
FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies 15
I57
Protein design from fold fragments
B. Hoecker
Max Planck Institute for Developmental Biology, Tu
¨
bingen,
Germany
Protein design can learn from mechanism observed in natural
evolution. We have applied the concept of modular assembly and
fragment recruitment to construct new well-folded proteins. By
combination of parts from two differently folded proteins, a
flavodoxin-like and a (ba)
8
-barrel protein, we were able to build
new ba-barrel. The crystal structure of one of the chimeric pro-
teins showed that the fragments retain their structures within the
new context. It further revealed an unexpected b-strand that is

important for the integrity of the fold. In order to construct a
stable protein without this strand we used computational design
to optimize the interfaces of the two fragments and to stabilize
the fold. Our approach illustrates how different protein folds
could have evolved from each other and developed anew. It fur-
ther shows the potential of this concept for the design of new
proteins from folding fragments where each fragment contributes
its own properties, e.g. ligand binding sites. Furthermore, we
want to use computational tools to design enzyme function and
ligand specificity into our proteins. Therefore, we performed a
structural analysis of a number of published receptors in order to
learn in molecular detail how precise the predictions of available
programs are. Our in-depth analysis did not confirm ligand bind-
ing, various methods showed no indication for binding as had
been previously described. Thus, although the computation of
side-chain conformations appears to be working, the design pro-
tocol for ligand-protein interactions has to be revisited and
improved.
I58
Abstract missing, please view Abstract Addendum
I59
Imaging HIV transfer between T cells with
optical superresolution
T. Huser
1
, G. McNerney
2
,W.Hu
¨
bner

3
, B. Dale
4
, D. Thompson
2
and B. Chen
4
1
Center for Biophotonics and Department of Internal Medicine,
University of California, Davis, Sacramento, CA, USA,
2
Center
for Biophotonics, University of California, Davis, Sacramento, CA,
USA,
3
European Molecular Biology Laboratory, Heidelberg,
Germany,
4
Department of Medicine, Division of Infectious
Diseases, Mount Sinai School of Medicine, New York, NY, USA
Live cell imaging with molecularly specific contrast is a particular
strength of optical microscopy, permitting the dynamic observa-
tion of events at the single cell level -either within a cell or inter-
actions between cells. This provides highly quantitative
biophysical data e.g. to unravel the systems biology of HIV-1
transfer between cells. Most live cell imaging experiments make
use of the high specificity provided by labeling cellular compo-
nents with fluorescent dyes and fluorescent proteins. Here, we
will present our latest data on tracking HIV-1 transfer between
cells by conducting 4D live cell fluorescence microscopy with a

replication-competent clone of the virus. A promising addition to
4D live cell microscopy comes in the form of holographic optical
tweezers, which enable us to actively manipulate infected and
uninfected cells within the field of view of the microscope and
which allow us to test the parameters affecting the initiation of
the virological synapse between cells. Another extension to con-
ventional fluorescence microscopy, 3D structured illumination
super-resolution microscopy, further allows us to dissect the spa-
tial structure of these adhesive synapses between cells. Combined,
these biophysical imaging and manipulation tools permit us to
exercise great control – temporally and spatially – over the events
preceding and following the transfer of viral material from
infected to uninfected cells and determine the nature of the trans-
fer mechanism.
I60
Regulation of epithelial cell migration,
invasion and EMT
J. Ivaska
University of Turku, Turku, Finland
One of the main interests in our laboratory is to understand the
co-operation between integrins and the cytoskeleton in regulation
of cells invasion. Epithelial-to-mesenchymal transition (EMT) is
a critical event in the progression towards cancer metastasis. The
intermediate filament protein vimentin is an important marker of
EMT and a requisite regulator of mesenchymal cell migration.
Past work in the laboratory has shown a role for vimentin in the
regulation of integrin traffic and cell migration. More recently,
we have identified that EMT induced by transcription factors is
critically dependent on upregulation of vimentin. In epithelial
sheets cells are normally bound to each other and to the extracel-

lular matrix to form ordered structures. In intercellular adhesions
tight junctions are essential for the mechanical strength and bar-
rier function of the epithelium. Integrin mediated cell-matrix
adhesions, on the other hand, link the extracellular matrix to the
cellular cytoskeleton. We have recently shown that migration of
lung cancer cells is controlled by a novel, spatially restricted a5
integrin-ZO-1 interaction, which is dynamically regulated by
PKCe, triggers persistent migration of these cells. This maybe
clinically relevant as loss of a tight junction protein, zonula oc-
cludens-1 (ZO-1), from intercellular adhesions, upregulation of
a5b1 integrin and activation of PKCe have all been show to cor-
relate with increased invasion and poor prognosis in lung cancer.
I61
Abstract missing, please view Abstract Addendum
I62
Implication of extracellular matrix in growth
and metastatic potential of human breast
cancer cells to bone: the role of zoledronate
N. Karamanos
1
,P.Dedes
1
,A.Tsonis
1
,C.Gialeli
1
and D. Kletsas
2
1
Lab of Biochemistry/Department of Chemistry, University of

Patras, Patras, Greece,
2
Lab of cell proliferation and aging/
N.C.S.R. ‘Demokritos’, Institute of Biology, Athens, Greece
Extracellular matrix (ECM) components such as proteoglycans,
metalloproteases and integrins contribute to growth and meta-
static potential of cancer cells as they are the barriers to tumor
cell migration and invasion. Breast cancer preferentially metas-
tasizes to bone, inducing locally increased bone resorption by
osteoclasts. Bisphosphonates (BPs) are analogues of PPi that
inhibit the function of the mevalonate pathway. BPs bind to
bone hydroxyapatite and are then released and internalized by
bone-resorbing osteoclasts. The effect of the BP zoledronate
(ZA) on key ECM molecules that promote tumourigenesis, cell
proliferation and metastasis was studied in breast cancer cells.
Furthermore, the effects of zoledronate (ZA) on the activation
of osteoclasts, premature osteoclasts incubated on bone particles
Abstracts Invited speakers
16 FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies
with the appropriate growth factors and co-cultures with breast
cancer cells were also evaluated. ZA effectively inhibits cell pro-
liferation, in vitro invasion and migration of cancer cells. Adhe-
sion of cancer cells to various matrices was reduced after
pretreatment with ZA and the levels of syndecan-1, gelatinases
and integrins amb3 and amb5 were significantly decreased.
Finally, the activation of pre-osteoclasts was inhibited in a dose
dependent mode either the activation was initiated by growth
factors either by co-culturing with breast cancer cells. The
results demonstrate that ZA may be a powerful therapeutic
agent in preventing the growth of breast cancer cells and their

invasion in bone.
I63
Systems biology of ageing and longevity
T. Kirkwood
Newcastle University, Institute for Ageing and Health, Newcastle
upon Tyne, UK
A longstanding barrier to progress in ageing research has been
the coexistence of multiple, seemingly competing hypotheses
about causal mechanisms. Each mechanism tends to be partially
supported by data indicating that it has a role in the overall cel-
lular and molecular pathways underlying the ageing process.
However, the magnitude of this role is usually modest. The sys-
tems biology approach combines (i) data-driven modeling, often
using the large volumes of data generated by functional genomics
technologies, and (ii) hypothesis-driven experimental studies to
investigate causal pathways and identify their parameter values
in an unusually quantitative manner, which enables the contribu-
tions of individual mechanisms and their interactions to be better
understood and it allows for the design of experiments explicitly
designed to test the complex predictions arising from such mod-
els. Examples of the success of the systems biology approach in
unraveling the complexity of ageing can be seen in recent studies
on cell replicative senescence, mitochondrial dysfunction and
breakdown in protein homeostasis. An important challenge also
exists in connecting the network of (random) damage-driven
proximate mechanisms of ageing with the higher level (genetically
specified) signalling pathways that influence longevity. This con-
nection is informed by actions of natural selection on the deter-
minants of ageing and longevity.
I64

Abstract missing, please view Abstract Addendum
I65
Molecular Design – from proteins to networks,
coupling computation and experiment
T. Kortemme
Department of Bioengineering and Therapeutic Sciences &
California Institute for Quantitative Biosciences, University of
California San Francisco (UCSF), San Francisco, CA, USA
We aim to link the details of interactions between biological mac-
romolecules to their in vivo functions within complex cellular net-
works, with two complementary motivations: to advance
quantitative understanding of biological processes and to enable
systematic cellular engineering. Towards these long-term goals,
my group pursues research projects at the all-atom, molecular
and systems-scale. I will describe our progress in two of these
areas: We develop foundational atom-level computational meth-
ods to predict and design protein structures and protein interac-
tions [1]. Most recently, we have shown that a new approach,
borrowing mathematical formulations from the field of robotics,
predicts the conformations of variable segments in proteins high
accuracy [2]. Applications include coupling this new method with
protein design to characterize fundamental relationships between
protein structure, sequence, dynamics and function of proteins
and to engineer proteins with new and useful functions. We com-
bine computation and experiment to dissect protein networks.
We systematically modulate protein interactions and determine
genome-wide changes in quantitative genetic interaction patterns
resulting from these perturbations, with the goal to map the sys-
tem-level functions of specific interactions in model organisms at
single-residue resolution.

References:
1. Mandell DJ & Kortemme T. Computer-aided design of functional
protein interactions. Nat Chem Biol 2009; 5:797–807.
2. Mandell DJ, Coutsias EA & Kortemme T. Sub-Angstrom Accu-
racy in Protein Loop Reconstruction by Robotics-Inspired Confor-
mational Sampling. Nature Methods 2009; 6:551–55 2.
I66
Hypothesis: Molecular basis of cholera blood
group dependence and implications for a
world characterized by climate change
A
˚
. Holmner
1
, A. Mackenzie
2
and U. Krengel
2
1
Department of Biomedical Engineering & Informatics,
Va
¨
sterbotten County Council, Umea
˚
, Sweden,
2
Department of
Chemistry, University of Oslo, Oslo, Norway
The severity of many infectious diseases, of bacterial or viral ori-
gins, correlates with the blood group of the infected individuals,

and cholera – the most severe diarrhoeal disease – serves as a
classic example of such blood-group-dependent diseases. Here we
review existing epidemiological, clinical and molecular data on
the subject and derive a hypothesis as to the molecular origins of
blood group dependence of cholera infections. The importance of
these investigations cannot be underestimated in the light of cli-
mate change. In response to increases in temperature, sea level
and precipitation variability in already exposed regions of the
world, water-borne diseases are likely to become an increasing
threat. The identification of individuals at particular risk for
severe disease will hence become increasingly important for crisis
minimization. Furthermore, understanding of the molecular basis
of blood group dependence may pave the way to the develop-
ment of novel intervention tools tailor-made for those groups of
the population that are at highest risk.
I67
Assessing the importance of individual players
in biochemical networks in a global way
U. Kummer
1
, S. Sahle
1
, S. Hoops
2
and P. Mendes
3
1
University of Heidelberg, Heidelberg, Germany,
2
Virginia Tech,

Blacksburg, VA, USA,
3
University of Manchester, Manchester,
UK
Modeling the dynamics of biochemical systems has become as
essential and integral part of systems biology. The rational
behind this fact is that a true understanding of biochemical net-
works must eventually result in valid predictions about the
behaviour of these networks in the living cell. In most cases, such
predictions will only be possible in the context of concise descrip-
tions of the networks which allow an analytical analysis. Mathe-
matical descriptions are doubtless concise and allow for a
multitude of ways to analyse their behaviour. However, in
Invited speakers Abstracts
FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies 17
general, a lot of precise experimental data is needed to build a
quantitative, computational model with predictive power. Since
such data is often unavailable, it is crucial to assess the impor-
tance of each process/player for the network dynamics to be able
to deal with rough estimates wherever possible and to specifically
ask for exact measurements wherever really needed. Such ques-
tions are often answered by a method called sensitivity analysis.
With this method, the impact of e.g. a specific kinetic parameter
on the dynamics of the network can be investigated. However,
often, this is only done in a local manner, for one specific set of
parameters. Not knowing many of them asks for more global
approaches. After a general introduction to the topic, this talk
will introduce some global ways to tackle the above questions
together with their limitations.
I68

Applications of cell-penetrating peptides in
oligonucleotide delivery
U. Langel
Stockholm University, Neurochemistry, Stockholm, Sweden
Recent achievements and our recent data are summarized on
improved drug delivery by Cell-Penetrating Peptides (CPPs) on
the variety of different cargoes including antisense and siRNA
oligonucleotides, peptides and proteins. These cargoes are applied
to regulate gene expression in live cells and in vivo, holding great
promise in further drug development. Some recent references fol-
low [1–3].
References:
1. Cell-Penetrating Peptides, 2nd Edition (Ed. Langel, U
¨
). CRC Press
Boca Raton, London, New York, Washington DC, 2006.
2. Ma
¨
e M, et al. A stearylated CPP for delivery of splice correcting
oligonucleotides using a non-covalent co-incubation strategy. J
Contr Release 2009; 134: 221–227.
3. Fossat P, et al. Knockdown of L calcium channel subtypes: differ-
ential effects in neuropathic pain. J Neurosci 2010; 30(3): 1073–
1085
I69
Metabolism and inflammation in human
adipose tissue
D. Langin
Institut de Me
´

decine Mole
´
culaire de Rangueil (I2MR), Inserm,
Universite
´
Paul Sabatier, CHU Toulouse, Toulouse, France
Understanding of the mechanisms controlling adipose tissue biol-
ogy is essential for the elucidation of the etiology of obesity com-
plications. Recently, progress has been made in several avenues
relating to human adipose tissue. Our knowledge of lipolysis (i.e.
the hydrolysis of triglycerides into fatty acids) has grown with
the identification of new lipases, cofactors and interactions
between proteins and lipids. The respective importance of adipose
triglyceride lipase, its co-factor CGI-58 and hormone-sensitive
lipase has been assessed in human fat cells. Through expression
or activation of transcriptional modulators such as PGC-1alpha
and PPARalpha, we have also provided evidence that human
mature white adipocytes can be converted into brown fat-like
adipocytes, and therefore that the typical fatty acid storage phe-
notype of white fat cell can be altered towards a fat utilization
phenotype. Human obesity is now known to be associated with
adipose tissue inflammation. Investigating subcutaneous and visc-
eral adipose tissues of groups of subjects with different obesity
and metabolic status revealed an opposite pattern between adipo-
cytes and macrophages. In lean subjects, genes related to adipo-
cyte metabolic pathways are highly expressed whereas genes of
macrophage inflammatory pathways show low expression. An
opposite pattern is seen in obese patients with metabolic syn-
drome suggesting a crosstalk between adipocytes and macro-
phages within adipose tissues which is related to the extent of fat

mass and the development of insulin resistance. These data open
up new opportunities for the development of drugs for obesity
metabolic and cardiovascular complications.
I70
Regulation of mtDNA expression in disease
and ageing
N G. Larsson
Max Planck Institute for Biology of Ageing, Cologne, Germany
The main task of the mitochondrial network is to convert food
into the cell’s main energy currency, adenosine tri-phosphate,
ATP. This is accomplished by the process of oxidative phosphor-
ylation. The mitochondrial function is fully dependent upon pro-
teins encoded by the mitochondrial DNA, mtDNA. Point
mutations and deletions of mtDNA accumulate in a variety of
tissues during ageing in humans, monkeys and rodents. To study
the link between mtDNA mutations, mitochondrial dysfunction
and ageing, we generated mice expressing an error-prone form of
the catalytic subunit of the mtDNA polymerase, PolgA. These
mice have an mtDNA mutator phenotype with a 3-fold to 5-fold
increase in the levels of point mutations, as well as increased
amounts of deleted mtDNA. This increase in somatic mtDNA
mutations is associated with reduced lifespan and premature
onset of ageing-related phenotypes such as weight loss, reduced
subcutaneous fat, alopecia, kyphosis, osteoporosis, anaemia,
reduced fertility and heart enlargement. We have used the
mtDNA mutator mice to derive mouse lines harboring high levels
of a few mtDNA point mutations. Such mice are valuable model
systems to study the role of individual mtDNA in disease and
ageing. Age-associated, acquired mutations of mtDNA often
undego clonal expansion to create a mosaic pattern of respiratory

chain deficiency. We have experimental studied the role of such
mutations by inactivating respiratory chain function in individual
cell types and by creating mouse chimeras.
I71
Cell wall and extracellular matrix in the human
opportunistic fungus Aspergillus fumigatus
J-P. Latge
Institut Pasteur, Paris, France
A major characteristic of all yeast and moulds is the presence of
a cell wall that surrounds the cell of these eukaryotic microorgan-
isms. The cell wall has both protective and aggressive functions
in fungal life. Protective, since it is the first barrier in contact
with the environment most often hostile encountered by the fun-
gus. Aggressive since all exchanges between the fungal cell and
its environment rely upon a functional and permeable cell wall.
The central skeleton of the fungal cell wall of the human oppor-
tunistic pathogen Aspergillus fumigatus is composed of chitin
and beta 1,3 glucans. They are separately synthesized as linear
polysaccharides by the polysaccharide synthases at the plasma
membrane and extruded in the cell wall space where they are
modified and cross-linked to other polymers. In addition to this
fibrillar skeleton, amorphous polysaccharides that are alpha 1,3
glucan and galactosaminogalactan play the role of cement for the
wall structure. These polymers are also the major component of
the extracellular matrix which glues together the hyphae of a
colonial mat in vitro under aerial conditions and in vivo and pro-
tects the fungus against external stresses.
Abstracts Invited speakers
18 FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies
Our current knowledge of the three-dimensional structural orga-

nisation of all these constitutive cell wall polysaccharides in A.
fumigatus and the enzymes that are responsible for their synthe-
sis, branching and cross-linking will be presented. I will also
emphasize the fact the cell wall cannot be considered any longer
as an inert exoskeleton, since its structure continuously evolves in
response to the environment and during cell cycle.
I72
Perturbing genes and predicting the outcome
B. Lehner
Centre for Genomic Regulation, EMBL-CRG Systems Biology,
Spain
Our aim is to understand and predict when perturbations are
harmful, and when they are not. This provides a framework for
studying evolution and also genetic disease. Recently in collabo-
ration with the Marcotte, Lee and Fraser labs we have shown
how integrated functional networks can be used to globally pre-
dict the effects of loss of function mutations [1] and also how
mutations in multiple genes combine to cause synthetic pheno-
typic change [2,3]. Further, we have investigated the mechanisms
that cause increased gene expression to be harmful and how det-
rimental dosage increases are physiologically avoided [4] . Many
mutations have little apparent effect because of genetic compen-
sation (‘redundancy’), and we aim to understand how compensa-
tion evolves [5,6]. As part of this we are investigating the general
principle that robustness to genetic and non-genetic perturbations
is coupled, depending on the same gene products [7,8].
References:
1. Lee et al. Nat Genet 2008.
2. Lee et al. Genome Res in press.
3. Lehner et al. Nat Genet 2006.

4. Vavouri et al. Cell 2009.
5. Tischler et al. Nat Genet 2008.
6. Vavouri et al. Trend Genet 2008.
7. Lehner. Mol Syst Biol 2008.
8. Lehner. PloS One 2010.
I73
Please view Plenary lectures
I74
Real-time microscopy and high-content
imaging as essential tools to dissect dynamics
of the DNA damage response
C. Lukas
Danish Cancer Society, Copenhagen, Denmark
Stochastic or clastogen-induced chromosomal breakage poses
extreme demands on coordinating DNA repair with vital cellular
processes such as gene expression, DNA replication or chromo-
somal segregation. To counteract deleterious effects of DNA
damage, cells activate genome surveillance pathways. These
molecular pathways operate both directly at damaged chromo-
somes and in the undamaged parts of the nucleus, and their
activity is constantly adjusted by signals from various DNA
intermediates as well as modified histones and histone-binding
proteins. Failure to coordinate these processes undermines gen-
ome integrity and may lead to fatal diseases including cancer.
Our laboratory is interested in how genome surveillance path-
ways operate on a molecular level and how are these processes
dynamically organised within the three dimensional space of the
nucleus. In my presentation I will discuss our approaches to
monitor in real time the intra-nuclear protein redistribution after
DNA damage, and provide evidence that a simple timing of pro-

tein assembly at and around the DNA breaks provides important
and sometimes unexpected insights into the physiology of gen-
ome surveillance. In addition, I will summarize the results from
our recent high-content microscopy screening projects and show
that it is a powerful tool to uncover hitherto unknown genome
caretakers involved in DNA repair and in protecting intrinsically
fragile regions of mammalian chromosomes.
I75
Intense parallel sequencing and high
sensitivity identification of mutations,
transcriptomes and genomes
J. Lundeberg
KTH, Royal Institute of Technology, Div Gene Technology,
Stockholm, Sweden
Advancements in the field of DNA sequencing are changing the
scientific horizon and promising an era of personalized medicine
for elevated human health. Although platforms are improving at
the rate of Moore’s Law, thereby reducing the sequencing costs
by a factor of two or three each year, we find ourselves at a
point in history where individual genomes are starting to appear.
Current parallel, state-of-the-art systems are providing signifi-
cantly improved throughput over Sanger systems with massive
deciphering of genetic sequences shedding light on novel biologi-
cal functions and phenotypic differences. At the new Science for
Life Laboratory, Stockholm that hosts the national center for
massive DNA sequencing (SNISS) we have improved the sample
preparation for the major next generation sequencing platforms
for more efficient and sensitive use of the technology. This lecture
will discuss some of these improvements and also demonstrate
the use of the platform to perform molecular archeology of stem

cell compartments in human epidermal tissue, correlation of tran-
scriptomes with proteomes, sequence capture of genomic regions
and analysis of genomes.
I76
The quest for molecular quasi-species in
ligand-activity space and its application to
directed enzyme evolution
B. Mannervik and A. Runarsdottir
Uppsala University, Uppsala, Sweden
We propose that the proper evolving unit in enzyme evolution is
not a single ‘fittest molecule’, but a cluster of related variants
denoted a ‘quasi-species’. A distribution of variants provides
genetic variability and thereby reduces the risk of inbreeding and
evolutionary dead ends. In our perspective, the ambient ligand
matrix governs the selection of novel enzymes with valuable func-
tional properties in evolution. Different matrices of substrates or
activity modulators will lead to different selection criteria and
divergent evolutionary trajectories. We provide examples from
our investigations of glutathione transferases illustrating the
interplay between libraries of enzyme variants and ligand matri-
ces in the emergence of quasi-species. In the directed evolution of
enzymes for novel functions, the ligand matrix is crucial to the
outcome of the search for novel activities. In this sense the exper-
imental system is similar to the biological environment in govern-
ing the evolution of enzymes.
Reference:
1. Emre
´
n LO, Kurtovic S, Runarsdottir A, Larsson A-K and Man-
nervik B. Functionally diverging molecular quasi-species evolve by

crossing two enzymes. Proc Natl Acad Sci U S A 2006; 103:
10866–10870.
Invited speakers Abstracts
FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies 19
I77
The pre-B cell receptor checkpoint
L. Martensson
Medicine, Gothenburg, Sweden
B-lymphocytes (cells) are central to the immune system in their
role as antibody producing cells. Antibodies protect by binding
to unique structures on pathogens, which leads to the formation
of immune complexes that are disposed off by the body. Each B
cell synthesises a unique antibody due to a process in which gene
segments are randomly recombined. This process, however, also
leads to the synthesis of autoantibodies, i.e. antibodies recognis-
ing self. To ensure the generation of a diverse antibody reper-
toire, recognising foreign but not self antigens, cells undergo
selection at several checkpoints. During early B cell development,
two major checkpoints are in place. At the first, cells have com-
pleted recombination of immunoglobulin heavy chain and, as this
is expressed, it assembles with surrogate light chain to form a
pre-B cell receptor (pre-BCR). At the second, cells have com-
pleted recombination also of immunoglobulin light chain and, as
this is expressed, it assembles with the pre-existing heavy chain
into a BCR, i.e. a membrane bound antibody. Here, we will
focus on the first, the pre-BCR checkpoint. We will discuss how
mutations affecting the pre-BCR prevents the development of
autoreactive B cells and hence, may underpin the development of
systemic autoimmunity.
I78

A functional role for protein kinase CK2 in the
Arabidopsis circadian system
S. Portoles and P. Mas
Centre for Research in Agricultural Genomics (CRAG);
Consortium CSIC-IRTA-UAB, Barcelona, Spain
Circadian rhythms are oscillations in biological processes with a
period of 24 h. These rhythms persist under constant environ-
mental conditions indicating that are generated by a self-sus-
tained endogenous oscillator known as circadian clock. In
Arabidopsis, clock genes and their protein products operate via
auto-regulatory feedback loops that promote the rhythmic oscil-
lations in metabolic and physiologic activities. An increasing
body of evidence also points towards a key role of post-transla-
tional regulation of clock components as an essential mecha-
nism of circadian function. In our studies, we identified CKB4,
a CK2 regulatory subunit, as a component of the Arabidopsis
circadian system. Our findings show that the phosphorylated
isoforms of CKB4 are the preferred substrate for ubiquitination
and degradation by the proteasome pathway. We provide evi-
dence of the involvement of the biological clock in the circa-
dian regulation of CKB4 protein abundance, which itself is
important for an accurate control of circadian period by the
clock. By using plants over-expressing CKB4, we found circa-
dian phenotypes that lead to clock dissonance with the environ-
ment and reduced plant sensitivity to day length. Furthermore,
the use of chromatin immunoprecipitation assays together with
pharmacological inhibitors and a CK2 dominant negative
mutant revealed that the phosphorylation activity of CK2
affects the binding of CCA1 (CIRCADIAN CLOCK ASSOCI-
ATED 1) to its target promoters. We propose that CK2 antag-

onizes CCA1 regulatory activity to modulate the transcriptional
network controlling morning and evening phase-specific expres-
sion in Arabidopsis.
I79
Abstract missing, please view Abstract Addendum
I80
Abstract missing, please view Abstract Addendum
I81
Trans-splicing is a regulated and is master
regulator of trypanosome gene expression
S. Michaeli
Faculty of Life Sciences, Bar-Ilan University, Ramat-Gan, Israel
Trypanosomatids are ancient eukryotes and the causative agents
of major parastic dieases Trypaansome gene expression is regu-
lated mainly post-transcriptionally. mRNAs are formed by a
concerted action of trans-splicing and polyadenylation. In try-
pnosomes, all mRNAs are tarns-spliced. In this process, a
spliced leader (SL) is donated to all mRNAs from a small
RNA, the SL RNA. Two unique mechanisms discovered in our
group will be described; SLS or spliced leader RNA silencing
and SLD-an SL RNA discard mechanism. SLS was discovered
as unique stress induced mechanism that is induced under ER
stress elicited either by blocking of entry of proteins to the ER
or by ER stressors. SLS transmits a signal to the nucleus to
shut-off SL RNA transcription and in one step the cell stops
splicing. We provide evidence that SLS is a programmed cell
death pathway that serves to quickly eliminate unfitted para-
sites from the population. SLD was discovered when cells were
depleted for the SL RNA core proteins. SLD results in the
accumulation of defective SL RNA that is discarded from the

nucleus to the cytoplasm and is eventually excreted from the
cell. SLD is a novel mechanism to discard defective RNA not
by degradation but via sequestration and excretion. Trans-splic-
ing is therefore the process that is able to change and respond
to physiological cues and hence must be major in regulating
the adaptation of these parasites during cycling between its
their two hosts
I82
Protein phosphorylation in bacteria
I. Mijakovic
AgroParisTech-INRA, MICALIS, Thiverval-Grignon, France
Protein phosphorylation is one of the most abundant post-
translational modifications present in all life forms. In Eukarya,
phosphorylation cascades of Ser/Thr and Tyr kinases constitute
one of the major signal transduction pathways, whereas in
Bacteria, this role was assigned to His/Asp kinases of the two-
component and phosphorelay systems. However, phosphoprote-
omics and other analytical approaches have recently revealed in
bacteria a complement of Ser/Thr and Tyr kinases and their
substrates that in many cases match that of the two component
systems. Currently, efforts are underway to chart these new
phosphorylation networks, assigning individual kinases to their
substrates, using state of the art proteomics and interactomics
tools. In parallel, studies are conducted to assay the functional
relevance of Ser/Thr and Tyr phosphorylation in bacteria. In
some cases, protein phosphorylation controls enzymatic activity
of the kinase substrates, in other cases their sub-cellular locali-
zation or half-life in the cytosol. A number of proteins phos-
phorylations are self-inflicted and the phosphorylated residues
Abstracts Invited speakers

20 FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies
participate in the reaction mechanism of the enzymes in ques-
tion. But there also seem to be phosphorylation events that do
not have any effect on the target proteins. Is that only because
we do not know where to look for effects, or could there be
an alternative explanation for these apparently ‘useless’ phos-
phorylations?
I83
The two terminal enzymes of denitrification:
nitric and nitrous oxide reductase
I. Moura
Requimte/CQFB, Dep.Quı
´
mica, FCT, UNL, Caparica,
Portugal
Denitrification, or dissimilative nitrate reduction, is an anaerobic
process used by some bacteria for energy generation. This process
is important in many aspects, but its environmental implications
have been given particular relevance. Nitrate accumulation and
release of nitrous oxide in the atmosphere due to excess use of fer-
tilizers in agriculture are examples of two environmental problems
where denitrification plays a central role. The reduction of nitrate
to nitrogen gas is accomplished by four different types of metal-
loenzymes in four simple steps. In this process, the nitrate mole-
cule is reduced to molecular di-nitrogen in a series of reactions:
2NO
3
-
fi 2NO
2

-
fi 2NO fi N
2
O fi N
2
We present a concise updated review of the bioinorganic aspects
of denitrification with emphasis on structural and mechanistic
aspects of the relevant enzymes involved in the two last steps of
this complex pathway. Biochemical and spectroscopic studies of
the two terminal enzymes will be discussed. Nitric oxide reductase
(NOR), a membrane enzyme contains a catalytic center composed
by a non-heme iron coupled to a b type heme. The crystal struc-
ture of N
2
OR was solved to a resolution of 2.4 A°. This enzyme
contains one binuclear (CuA) and a tetranuclear copper center
(CuZ), an unusual structure (catalytic site). CuZ center is a new
type of cluster, in which four copper ions are coordinated by seven
histidine residues. A model is proposed for the binding of N
2
O
binds to this center. EPR, X-ray and DFT have been crucial unre-
vealing unique structural aspects. Most of the reactions involve
O-atom abstraction and mechanistic proposals will be presented.
I84
Structural insights into eukaryotic aquaporin
regulation
S. To
¨
rnroth-Horsefield

1
, K. Hedfalk
1
, G. Fischer
1
, K. Lindkvist-
Petersson
2
and R. Neutze
1
1
Chemistry, University of Gothenburg, Gothenburg, Sweden,
2
Cell
and Molecular Biology, University of Gothenburg, Gothenburg,
Sweden
Aquaporin mediated water transport across cellular membranes
is an ancient, ubiquitous mechanism within cell biology. This
family of integral membrane proteins includes both water selec-
tive pores (aquaporins) and transport facilitators of other small
molecules such as glycerol and urea (aquaglyceroporins). Eukary-
otic aquaporins are frequently regulated post-translationally by
gating, whereby the rate of flux though the channel is controlled,
or by trafficking, whereby aquaporins are shuttled from intracel-
lular storage vesicles to the plasma membrane. A number of
high-resolution X-ray structures of eukaryotic aquaporins have
recently been reported and the new structural insights into gating
and trafficking that emerged from these studies are described.
Basic structural themes reoccur, illustrating how the problem of
regulation in diverse contexts builds upon a limited set of possi-

ble solutions.
I85
Abstract missing, please view Abstract Addendum
I86
Metabolism of filamentous fungi at the
genome level
J. Nielsen
Chalmers University of Technology, Gothenburg, Sweden
Filamentous fungi are used extensively in the fermentation indus-
try for the production of antibiotics, industrial enzymes and
organic acids. In connection with further development of biopro-
cesses for the production of fuels and chemicals these fungi are
interesting versatile cell factories as they tolerates low pH, can
utilize a wide range of carbon sources and has relatively high
conversion rates. With the availability of the genome sequence
for several different fungi it has become possible to query the
metabolic capabilities of these microorganisms at the genome-
level. Using a bottom-up approach using genomic information
together with information from databases, research papers and
books, we have reconstructed the metabolic networks of A. nidu-
lans, A. niger and A. oryzae. In connection with the metabolic
network reconstruction process we further improved the genome
annotation. The metabolic networks were used as scaffolds for
identification of co-regulatory modules in response to different
perturbations, e.g. changes in the carbon source and this has
allowed us to analyze differences in the metabolic responses
between the three species. The metabolic networks have also been
converted into so-called genome-scale metabolic models that can
be used for identification of metabolic engineering targets. Using
the genome-scale metabolic models for these three species we

developed tools for semi-automatic reconstruction of a genome-
scale metabolic model for the filamentous fungus Penicillium
chrysogenum, that is used for penicillin production. The model
will be presented and novel insight gained into the metabolism of
this industrially important fungus will be discussed.
I87
Abstract missing, please view Abstract Addendum
I88
Phosphatidylinositol 4-phosphate controls
both membrane recruitment and a regulatory
switch of the Rab GEF, Sec2p
E. Mizuno-Yamasaki
1
, J. Coleman
2
, M. Medkova
3
and P. Novick
1
1
Cellular and Molecular Medicine, University of California San
Diego, San Diego, CA, USA,
2
Cell Biology, Yale University
School of Medicine, New Haven, CT, USA,
3
RainDance Technolo-
gies, Lexington, MA, USA
Sec2p is the guanine nucleotide exchange factor (GEF) that acti-
vates the Rab GTPase Sec4p on secretory vesicles. Sec2p also

binds a Rab acting earlier in the secretory pathway, Ypt32p-GTP,
forming a RabGEF cascade. Ypt32p and the Sec4p effector
Sec15p (a component of the exocyst complex) compete for bind-
ing to Sec2p. Indeed Ypt32p initially recruits Sec2p, but subse-
quently allows a handoff of active Sec2p/Sec4p to Sec15p.
Intriguingly, Golgi-associated phosphatidylinositol 4-phosphate
(PI4P) works together with Ypt32-GTP in this context. PI4P
inhibits Sec2p-Sec15p interactions, promoting recruitment of
Sec2p by Ypt32p as secretory vesicles form. However, PI4P levels
appear to decline as vesicles reach secretory sites, allowing Sec15p
to replace Ypt32p as vesicles mature. In this way, the regulation
of PI4P levels may switch Sec2p/Sec4p function during vesicle
Invited speakers Abstracts
FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies 21
maturation, from a RabGEF recruitment cascade involving
Ypt32p, to an effector positive feedback loop involving Sec15p.
I89
Unequal inheritance of protein damage during
yeast cytokinesis
T. Nystrom
University of Gothenburg, Gothenburg, Sweden
The paradigm sirtuin, Sir2p, of budding yeast is required for reju-
venation of yeast progeny. This rejuvenation includes the retention
of damaged and aggregated proteins in mother cells. Since many
imperative pathways are redundant, we hypothesized that machin-
eries involved in segregating cellular damage could be identified by
establishing the global genetic interaction network of SIR2 using
synthetic genetic array (SGA) analysis. Using this approach, we
found unexpected genetic interactions with genes involved in estab-
lishing cell polarity, including members of the polarisome, the for-

min BNI1, and MYO2, encoding the class V myosin motor
protein. By visualizing Hsp104-containing aggregates we show that
the polarity machinery, including the tail of Myo2 required for
interaction with different cargoes, is essential for the retention of
protein aggregates during mitotic cytokinesis. In addition, we
found that this machinery can effectively clear the progeny from
damaged proteins during stress by a retrograde transport of protein
aggregates. Our findings are in accord with recent models hypothe-
sizing that polarity may have evolved to avoid clonal aging.
I90
The Escherichia coli replisome and use of
clamps to bypass replication barriers
M. O’Donnell, R. E. Georgescu and N. Y. Yao
Rockefeller University, HHMI, New York, NY, USA
Chromosomal replicases utilize circular sliding clamps for high-
processivity during replication. Sliding clamps not only bind the
chromosomal replicase, but they also function with all five DNA
polymerases in E. coli as well as other proteins for repair and
lesion bypass. This presentation focuses on the use of the sliding
clamp by the replicase in crossing barriers that are encountered
during replication. Studies have shown that the replicase, Pol III,
can rapidly hop from one sliding clamp to another clamp in
order to transfer to the multiple RNA primers synthesized during
lagging strand replication. However, recent studies indicate that
polymerase hopping among sliding clamps is of more general
use, and can allow the polymerase to bypass lesions and even to
circumvent a tightly bound RNA polymerase that it encounters
during replication. We will present studies that demonstrate the
how the replisome deals with collisions with RNA polymerase
transcribing either the leading or lagging strand template. We will

also present our findings on how different polymerases control
the use of the clamp to form alternative replisomes for DNA
damage avoidance and, when needed, to deal with lesions that
are encountered directly. Finally, we present single-molecule stud-
ies that examine replisome action during lagging strand synthesis
and the consequences for the leading strand polymerase.
I91
Reconstruction and multi-scale modeling of
lipid networks
M. Oresic
VTT Technical Research Centre of Finland, Espoo, Finland
Lipids are a highly diverse class of molecules with crucial roles in
cellular energy storage, structure and signaling. Lipid homeosta-
sis is fundamental to maintain health, and lipid defects are cen-
tral to the pathogenesis of important and devastating diseases.
The spatial complexity of lipid metabolism presents a challenge
for the study of the global lipidomic profiles. Studies of lipidomic
profile in the context of biochemical pathways may shed light on
processes behind the synthesis of or regulation by specific lipids,
but cannot address how these changes translate into lipid mem-
brane properties and affect cellular physiology. However, molecu-
lar modeling tools and large scale computing capacities are
becoming available which facilitate modeling of lipid bilayers in
the context of their composition and function, thus opening new
opportunities to interpret regulation and changes of global lipi-
dome also in the spatial and physiological context. In this presen-
tation, two recent applications will be described which combined
clinical research, lipidomics, and multi-scale lipid modelling: (i)
investigations of HDL structure and lipid spatial distribution in
subjects with high and low HDL cholesterol; leading to poten-

tially important insights into the HDL function, and (ii) investi-
gations of adipose tissue phospholipids in the context of acquired
obesity, using a twin study design.
I92
Controlling the rates of biochemical reactions
and signaling networks by shape-and-volume
changes
O. Orwar
Chalmers University of Technology, Gothenburg, Sweden
Organelles as well as surfactant nanostructures (nanotube net-
works) are nano-scale, pleiomorphic systems with a capacity for
shape changes. In both these systems, transport, mixing, and shape
changes can be achieved or very close to thermal energy levels. In
further contrast to macroscopic systems, mixing by diffusion is
extremely efficient, and the kinetics of embedded reactions can be
controlled by shape- and volume changes. The coupling between
shape changes, and chemical activity is often strong, and cases will
be presented where chemistry affect reactor geometry, where reac-
tor geometry affect chemistry, and cases where the two properties
feed back on each other in self-regulating systems. We will show
several non-intuitive and fascinating dynamic properties in a vari-
ety of such systems including front propagation in reaction-diffu-
sion networks consisting of nanotube-conjugated containers,
oscillatory behavior for reversible reactions in volume-fluctuating
systems, and filtering of chemical signals in small networks. Using
volume fluctuations in mitochondria as an example, we show that
the rate of product formation of an enzymatic reaction can be reg-
ulated by simple volume transitions. We also show that spatiotem-
poral properties of reaction-diffusion systems are extremely
sensitive to sudden changes in network topology, and that chemi-

cal reactions can be initiated or boosted in certain nodes as a func-
tion of connectivity. In addition, distant links may be used to
synchronize spatially separated parts of the network. Finally, we
demonstrate that product inhibition and correlation of autocata-
lytic reactions can be controlled by network wiring.
I93
Cargo recognition by the AP2 endocytic
adaptor complex
B. Kelly
1
, S. Hoening
2
, P. Evans
3
and D. Owen
1
1
University of Cambridge, CIMR, Cambridge, UK,
2
Depatrment
of Biochemistry, University of Cologne, Cologne, Germany,
3
MRC
LMB, Cambridge, UK
Clathrin mediated endocytosis from the cell’s limiting membrane
plays a vital role in many cellular processes including signalling,
Abstracts Invited speakers
22 FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies
homeostasis, nutrient uptake and in defining the interactions of
cells with its surrounding. It is also the route by which many

pathogens gain access to the cell. The AP2 adaptor complex (a/
b2/l2/r2 subunits) is the major clathrin adaptor in endocytic
clathrin-coated vesicles where it crosslinks the clathrin scaffold to
PtdIns4,5P2-containing plasma membrane and also to transmem-
brane protein cargo embedded in it. In the ‘locked’ and inactive
cytosolic form, the binding sites for the two major endocytic
cargo motifs, Yxx F on the C-terminal domain of l2 (C-l2) and
[ED]xxxL[LI] on r2, are blocked by different parts of b2 subunit.
In order to bind cargo, the complex undergoes a large conforma-
tional change in which the 30 kDa C-l2 becomes relocated to a
different face of the complex resulting in AP2’s four PtdIns4,5P2
binding sites and two endocytic motif binding sites becoming co-
planar. Another consequence of the conformational change is
that the b2 subunit is no longer able to block the cargo motif-
binding sites. Liposome-based SPR and fluorescence polarisation
anisotropy data shows that the structural rearrangement of AP2
is driven by its association with the planar, polyanionic,
PtdIns4,5P2-containing plasma membrane. This membrane-acti-
vated switching mechanism prevents inappropriate binding to
YxxF and [ED]xxxLL sequences on cytoplasmic proteins and
also ensures that AP2 only recognises motif-containing cargoes
when they are at the cell’s limiting membrane and not when they
are in other intracellular membranes.
I94
Please view Plenary lectures
I95
The meta-structure of the E. coli genome and
its genome-scale transcriptional regulatory
network
B. Palsson and B. K. Cho

UCSD, Bioengineering, La Jolla, USA
Regulation of gene expression is widely considered to be a core
issue in biology. Normally such studies have focused on a single
gene or a group of genes. Advances in ChIP-chip technology
now enable us to study this issue on a genome scale. We have
obtained close to a 100 ChIP-chip data sets for E. coli K-12
MG1655 that include; RNAP, the sigma factors, most of the
broad acting transcription factors (TFs), and the majority of the
nucleoid associated protein (NAPs). In addition we have
obtained tiling array data for expression profiling and transcrip-
tion start site (TSS) data from short read sequencing technology.
Degradation start sites (DSS) are obtained by a variation of the
TSS protocol. This data enables us to (i) formulate the metastru-
cure (that is comprised of the functional, structural and opera-
tional annotation) of the E. coli genome and (ii) to draft the
reconstruction of the transcriptional regulatory network (TRN)
in E. coli at a genome-scale.
I96
From understanding to designing enzymatic
networks
S. Billerbeck, M. Bujara, C. Hold, R. Pellaux and S. Panke
Department of Biosystems Science & Engineering, ETH Zurich,
Basel, Switzerland
Biotechnology plays an increasingly central part in the manufac-
turing of compounds in the pharmaceutical, chemical, and fuel
industry. The underlying biological research has moved beyond
the molecular reductionist dogma to a systems view. This has re-
inforced the notion that the reliable and predictable construction
of novel biocatalytic systems is at odds with the complexity of
biological systems and highlights the need for concepts that allow

reducing this complexity in an attempt to recruit engineering
approaches for systems design. At the ETH Bioprocess Labora-
tory, we concentrate on the rational engineering of in vitro multi-
enzyme reaction networks, in particular for the production of
natural and unnatural sugars and ultimately oligosaccharides.
Crucial questions on our way to truly designing such networks
are how to insulate efficient pathways from highly interconnected
networks, such as the central carbon metabolism, and how to
optimize these pathways in terms of dynamic behavior. We will
illustrate possible strategies using our efforts in multi-enzyme
production of building blocks for C-C-bond forming enzymes
and real-time analysis of in vitro metabolic networks.
I97
Abstract missing, please view Abstract Addendum
I98
Autoimmune Regulator: molecular
mechanisms of central immune tolerance
P. Peterson
University of Tartu, Tartu, Estonia
Negative selection of T cells in the thymus is necessary for
the maintenance of self tolerance. The Autoimmune Regulator
(AIRE) protein is expressed in thymic medullary epithelial cells,
where it promotes the ectopic expression of tissue-specific antigens
needed for efficient negative selection of developing thymocytes.
Mutations in AIRE cause APECED (autoimmune polyendocrine
candidiasis ectodermal dystrophy) disease, in which patients
develop several autoimmune diseases. Similarly, Aire deficient
mice have a defect in negative selection and several recent studies
have shown that mutations in Aire lead to the escape of self-reac-
tive T cells from the thymus, which consequently results in au-

toimmunity. We have studied the molecular basis of AIRE-
mediated transcriptional regulation and our data show that AIRE
is able to promote ectopic gene expression from chromatin associ-
ated with histone modifications characteristic to inactive genes.
I99
The mitochondrial machinery for import and
assembly of proteins
N. Pfanner
University of Freiburg, Freiburg, Germany
Mitochondria contain about 1000 different proteins. Ninety-nine
percent of the proteins are encoded by nuclear genes and are syn-
thesized on cytosolic polysomes. Targeting signals direct the pre-
cursor proteins to receptors on the mitochondrial surface that are
part of the translocase of the outer membrane (TOM complex).
After passage through the channel formed by the TOM complex,
the precursor proteins are sorted into the mitochondrial subcom-
partments. (i) Preproteins with cleavable amino-terminal prese-
quences are imported by the presequence translocase of the inner
membrane (TIM23 complex) and the presequence translocase-
associated motor (PAM) of the matrix. Processing peptidases
remove the presequences and generate the mature proteins. (ii)
The precursors of hydrophobic metabolite carriers are trans-
ported across the intermembrane space by the Tim9-Tim10 chap-
erone complex and are inserted by the carrier translocase of the
inner membrane (TIM22 complex). (iii) The mitochondrial inter-
membrane space assembly (MIA) machinery mediates the import
and oxidative folding of proteins with characteristic cysteine
Invited speakers Abstracts
FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies 23
motifs. (iv) The mitochondrial outer membrane has different

pathways for insertion of alpha-helical proteins and beta-barrel
proteins. The sorting and assembly machinery (SAM complex)
inserts beta-barrel proteins into the outer membrane.
I100
Accumulation of triglycerides in green
microalgae: a potential source for biodiesel
P. Uri, Z. Tatyana and W. Meira
Biological Chemistry, The Weizmann Institute of Science, Rehovot,
Israel
Microalgae are recognized as a promising future source for large-
scale biodiesel production. Certain strains of microalgae accumu-
late large amounts of oil (mainly triglycerides,TG) that can be
extracted and converted by a simple chemical process to biodie-
sel. Microalgae display very high growth rates and oil productiv-
ity surpassing all oil crop plants and they do not compete with
agricultural lands or water resources. The major drawbacks for
economical biodiesel production from microalgae are partly tech-
nological, including the high cost of algae cultivation and pro-
cessing, and partly biological, the difficulties in keeping stable
cultures for extended periods, the poor state of knowledge about
regulation of TG accumulation and the lack of efficient genetic
tools to manipulate most oil accumulating algae species. On-
going research in universities and commercial companies is aimed
to improve biomass productivity and oil levels in microalgae by
genetic manipulations that includes improving light utilization in
dense cultures by decreasing the light harvesting antenna size,
improvement of carbon assimilation at high CO
2
levels, upregula-
tion of rate-limiting enzymes in TG biosynthesis, identification of

the signaling mechanism that controls TG accumulation and
development of herbicide-resistant lines to defeat contaminations.
Our strategies to maximize oil productivity from microalgae are
to utilize salt-resistant fast-growing strains suitable for extended
out-door cultivation, enhancement of TG biosynthesis by exter-
nal intervention and attempts to convert starch accumulating to
TG accumulating strains by genetic engineering.
I101
Quantitative targeted proteomics for the
analysis of cellular networks
P. Picotti
ETH Zuerich, Institute of Molecular Systems Biology, Zuerich,
Switzerland
To study and model cellular networks’ e.g. metabolic or signal-
ling networks’ it is crucial to measure all the elements that consti-
tute them, which are often associated to a wide range of
molecular properties and cellular abundances. However, compre-
hensive measurements are technically difficult, even in the simple
organism yeast, especially at the proteome level. To overcome
the limitations of classical approaches we applied a targeted pro-
teomic workflow based on selected reaction monitoring (SRM) to
the analysis of yeast cellular networks. We demonstrated that
proteins spanning the whole range of abundance, between 1.3E6
and <50 copies/cell, could be detected by SRM in yeast total
proteome digests. We then applied SRM to the quantification of
proteins in a complete metabolic network in yeast grown under
conditions inducing radically different metabolic setups and in a
time-course of yeast cells transiting through a series of metabolic
phases. All the SRM assays developed were deposited to the
web-accessible MRMAtlas database, which supports the collec-

tion and dissemination of the assays. Finally, to overcome the
bottlenecks of SRM assay development, we introduced a method
based on the use of unpurified synthetic peptide libraries, that
allows for the high-throughput and low-cost generation of vali-
dated SRM assays (>100 SRM assays/h) for any set of proteins
or proteome of interest. The approach was used to develop a
complete set of SRM assays for all the ~6600 proteins that con-
stitute the proteome of Saccharomyces cerevisiae and to a first set
of 8500 human proteins. The power and bottlenecks of this
approach will be discussed.
I102
The homologous collagens XV and XVIII have
distinct physiological roles
T. Pihlajaniemi
Oulu Center for Cell-Matrix Research, Biocenter Oulu and Insti-
tute of Biomedicine, University of Oulu, Oulu, Finland
Collagens XV (ColXV) and XVIII (ColXVIII) are homologous,
multidomain molecules occurring widely in association with base-
ment membranes (BMs). In human, mutations in the COL15a1
gene have not yet been reported but in mouse lack of ColXV
leads to mild myopathy. Mutations in the COL18a1 gene result
in Knobloch syndrome, a rare disorder characterized by eye
abnormalities and occipitocele, and in mouse complete lack of
ColXVIII also leads to various eye defects and increased suscept-
ibility to hydrocephalus. Our recent studies show that ColXV
plays a role in the pericellular fibrillar collagen organisation
whereas ColXVIII is required for proper BM structure in several
tissues. Attempts to characterise the physiological roles of ColXV
revealed that its lack in mice leads to changes that mimic a pre-
stage of dilated cardiomyopathy and predispose to pathological

responses under cardiac stress. Moreover, lack of ColXV per-
turbs microvascular stability and hemodynamics. To study the
specific roles of ColXVIII variants derived from the two distinct
promoters of the corresponding gene, we generated mutant
mouse lines expressing exclusively the short form (Col18a1del3/
del3), or alternatively, the two longer variants of the molecule
(Col18a1del1,2/del1,2). Blood sample analysis revealed that lack
of ColXVIII in mice, and specifically its long variants, leads to
increased levels of circulating triglycerides and a 2-fold increase
in the VLDL triglyceride content. The talk will describe these
and other recent data regarding the distinct properties and phy-
siological roles of the two homologous collagens.
I103
Abstract missing, please view Abstract Addendum
I104
Substrate-binding proteins: a structural
classification and the mechanism of peptide
selection by the transport receptor OppA
B. Poolman
Department of Biochemistry, University of Groningen, Groningen,
Netherlands
Substrate-binding proteins (SBP) are associated with a wide vari-
ety of protein complexes. The proteins are part of ATP-binding
cassette transporters, ion gradient driven transporters, DNA-
binding proteins, as well as channels and receptors from both
pro- and eukaryotes. A wealth of structural and functional data
is available on SBPs, with over 120 unique entries in the Protein
Data Bank. Over a decade ago these proteins were divided into
three structural classes, but based on the currently available
wealth of structural data, we propose a new classification into

six clusters, based on features of their three-dimensional struc-
ture. The SBP oligopeptide-binding protein A (OppA) from
L. lactis was crystallized and its structure was solved in the
Abstracts Invited speakers
24 FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies
open-unliganded conformation, along with structures of the li-
ganded protein in both open- and closed conformations [1]. The
structures directly explain the phenomenal promiscuity of the
OppA protein, which binds peptides of an exceptionally wide
range of lengths (4–35 residues), with no apparent sequence pref-
erence [2]. OppA’s binding cavity has an enormous volume (com-
patible with very long peptides), does not constrain the position
of the N-and C-termini (compatible with peptides of different
length), and avoids sequence-specific side-chain interactions.
Unexpectedly, the peptide’s amino acid composition (but not the
exact sequence) appears to play a role in selection, with a prefer-
ence for proline-rich peptides containing isoleucine. We propose
a mechanism for peptide selection based on amino acid composi-
tion rather than sequence.
References:
1. Berntsson et al. EMBO J 2009; 28: 1332.
2. Detmers et al. PNAS USA 2000; 97: 12487.
I105
Control of gene expression by the Hog1
stress-activated protein kinase
E. de Nadal and F. Posas
Universitat Pompeu Fabra, Barcelona, Spain
Exposure of yeast cells to increases in extracellular osmolarity
results in the activation of the Hog1 MAP kinase. Activation of
this MAP kinase is required to survive under high osmolarity

conditions. Adaptation to osmostress requires the induction of a
large number of genes, which indicates the necessity to regulate
several aspects of the cell physiology. Induction of gene expres-
sion is highly dependent on the presence of the MAP kinase,
which suggests a key role for the HOG signaling pathway in the
regulation of gene expression in response to osmostress. In
response to stress, the MAPK controls several mechanisms
related to transcription initiation; phosphorylation of transcrip-
tion factors, chromatin modification and recruitment of PolII.
Recruitment of Hog1 to ORFs during elongation is also critical
for proper mRNA production in response to stress, actually, one
of the aspects controlled by Hog1 is the reorganization of Chro-
matin.
I106
How well evolved is the folding code?
S. Radford
Faculty of Biological Sciences, University of Leeds, IMCB, Leeds,
UK
Proteins fold on rugged energy landscapes in which partially
folded structures are populated. Whilst such species may form
essential stepping stones to the native state, others may allow
access to misfolding pathways. Key to understanding the parti-
tioning between folding and misfolding, therefore, is to elucidate
the nature of the folding energy landscape in atomistic detail
and to identify features of non-native species that favour correct
folding or promote aberrant folding. Such information is not
only of fundamental importance, but underpins our quest to
elucidate the mechanism of chaperone action and the events
that occur to tip the balance between folding and aggregation.
In this lecture I will describe how we have determined folding

landscapes in atomistic detail using a combination of experi-
ment and simulation. Combined with further directed evolution
experiments I will show how the conflicting forces of folding,
stability and function has resulted in the evolution of today’s
amino acid sequences that encode functional, yet only margin-
ally stable protein products.
I107
Please view Plenary lectures
I108
Computational enzymatic catalysis
M. J. Ramos
Chemistry and Biochemistry, Requimte, Faculty of Sciences,
University of Porto, Porto, Portugal
We present theoretical studies on the mechanisms of enzymatic
reactions. We have resorted to QM and hybrid QM/MM calcula-
tions to analyze the energetics of the established mechanisms, or
proposed alternative pathways, and evaluated their feasibility
according to the observed kinetics of the enzyme and other exist-
ing experimental data. Our studies [1–4] have been very successful
in reproducing the existing experimental data.
References:
1. Bras NF, Fernandes PA, Ramos MJ. QM/MM Studies on the
beta-Galactosidase Catalytic Mechanism: Hydrolysis and Transgly-
cosylation Reactions. J Chem Theory Comput 2010; 6: 421–433.
2. Sousa SF, Fernandes PA, Ramos MJ. The Search for the Mecha-
nism of the Reaction Catalyzed by Farnesyltransferase. Chem Eur
J 2009; 15: 4243–4247.
3. Dourado DFAR, Fernandes PA, Mannervik B, Ramos MJ. Gluta-
thione Transferase: New Model for Glutathio ne Activation. Chem
Eur J 2008; 14: 9591–9598.

4. Ramos MJ, Fernandes PA. Computational Enzymatic Catalysis.
Acc Chem Res 2008; 41: 689–698.
I109
Methodology development in directed
evolution
M. T. Reetz
Max-Planck-Institut fu
¨
r Kohlenforschung, Mu
¨
lheim an der Ruhr,
Germany
Directed evolution is a protein engineering technique which allows
the genetic manipulation of almost any property of a protein,
including the enhancement of thermostability, substrate accep-
tance, and stereoselectivity of enzymes as catalysts in organic
chemistry and biotechnology. Many different gene mutagenesis
methods and strategies have been used, and indeed laboratory evo-
lution is always ‘successful’. However, true success also considers
the question of efficacy, setting the stage for developing fast and
efficient techniques for probing protein sequence space. Our con-
tribution is Iterative Saturation Mutagenesis (ISM), which requires
only small mutant libraries and therefore considerably less screen-
ing effort. ISM studies in the author’s laboratory have been carried
out in the quest to enhance stereoselectivity, broaden substrate
scope and increase thermostability of enzymes, and other groups
have joined efforts in testing ISM in these and other areas of appli-
cation. The success of ISM has been traced to the absence of super-
fluous mutations and to strong cooperative effects operating
between point mutations and sets of point mutations along evolu-

tionary pathways. The scope and limitation of this protein engi-
neering method will be delineated.
References:
1. Reetz MT, Kahakeaw D, Sanchis J. Mol Bio Syst 2009; 5: 115–
122.
2. Reetz MT. J Org Chem 2009; 74: 5767–5778.
3. Reetz MT, Wu S. J Am Chem Soc 2009; 131: 15424–15432.
4. Wu S, Acevedo JP, Reetz MT. Proc Nat Acad Sci U S A
2010; 107: 2775–2780.
5. Reetz MT Angew. Review of ISM. Chem Int Ed 2010, in press.
Invited speakers Abstracts
FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies 25
I110
Protein-protein interactions in MAP kinase
mediated signaling networks
A. Remenyi
Biochemistry, Eotvos Lorand University, Budapest, Hungary
It is an intriguing problem how one can make up highly specific
signaling networks by using promiscuous components. Mitogen
activated protein kinases (MAPK) possess substrate binding
pockets with surprisingly low specificity. However, they use bin-
ary docking interactions that are independent of the enzyme’s the
active site and are mediated through linear sequence motifs from
binding partners. In addition, they also employ multivalent scaf-
fold proteins to build up signaling modules comprised of other
enzymes or substrates. We have probed the specificity of MAPK
docking interactions and characterized the associations of MAPK
module components to several signaling scaffolds biochemically.
Our results demonstrate how these interactions enable MAPKs
to operate specifically in the context of their signaling modules.

I111
TLR4 mediated skin carcinogenesis is
dependent on immune and radioresistant cells
D. Mittal
1
, F. Saccheri
1
,E.Ve
´
ne
´
reau
2
, T. Pusterla
2
,
M. E.Bianchi
2
and M. Rescigno
1
1
European Institute of Oncology, Milan, Italy,
2
Universit
Vita-Salute San Raffaele, Milan, Italy
Skin cancers are the most commonly diagnosed cancers. Under-
standing what are the factors contributing to skin tumor develop-
ment can be instrumental to identify preventive therapies. The
myeloid differentiation primary response gene (MyD)88, the
downstream adaptor protein of most Toll-like receptors (TLR),

has been shown to be involved in several mouse tumorigenesis
models. We show here that TLR4, but not TLR2 or TLR9, is
upstream of MyD88 in skin tumorigenesis. TLR4 triggering is
not dependent on LPS associated to skin-colonizing bacteria but
on the high mobility group box-1 protein (HMGB1), an endo-
genous ligand of TLR4. HMGB1 is released by necrotic kerati-
nocytes and is required for the recruitment of inflammatory cells
and for the initiation of inflammation. The expression of TLR4
on both bone marrow derived and radioresistant cells is necessary
for carcinogenesis. Consistently, a human tissue microarray ana-
lysis showed that melanoma and colon cancer display an over
expression of TLR4 and its downstream adaptor protein MyD88
within tumors. Together, our results suggest that the initial
release of HMGB1 triggers a TLR4-dependent inflammatory
response that leads to tumor development.
I112
Photosystem II at 2.9 A
˜
. resolution – Quinones,
lipids, channels and chloride ion
W. Saenger
1
, A. Guskov
1
, A. Gabdulkhakov
1
, M. Broser
2
,
J. Kern

2
and A. Zouni
2
1
Freie Universitaet Berlin, Chemie und Biochemie/Kristallographie,
Berlin, Germany,
2
Technische Universitaet Berlin, Max Volmer
Laboratorium, Berlin, Germany
Photosystem II (PSII) is a homodimeric protein-cofactor complex
acting as light-driven water:plastoquinone oxidoreductase and is
located in the photosynthetic thylakoid membrane of plants,
green algae and cyanobacteria. PSII oxidizes two water molecules
at the unique Mn4Ca cluster to molecular (atmospheric) oxygen,
4 protons and 4 electrons. The protons drive ATP synthetase,
and the electrons reduce plastoquinone (QB) to plastoquinol
(QBH2) that is exported and delivers the electrons (through the
cytochrome b6f complex) to photosystem I. Here the electrons
gain a high reducing potential and serve at NADP reductase to
generate NADPH that together with ATP reduces CO
2
to carbo-
hydrates in the Calvin cycle. The crystal structure of PSII from
Thermosynechococcus elongatus at 2.9-A
˜
. resolution allowed the
unambiguous assignment of all 20 protein subunits and complete
modeling of all 86 cofactors, among them 25 integral lipids, per
PSII monomer. The presence of a third plastoquinone QC and a
second plastoquinone-transfer channel, which were not observed

before, suggest mechanisms for plastoquinol-plastoquinone
exchange, and we calculated possible water or dioxygen and pro-
ton channels. Putative oxygen positions obtained from Xenon
derivative crystals indicate a role for lipids in oxygen diffusion to
the cytoplasmic side of PSII. The chloride position suggests a
role in proton-transfer reactions because it is bound through a
putative water molecule to the Mn4Ca cluster and is close to two
possible proton transfer channels.
I113
Experiencing research through creative design
and ownership of laboratory practicals
J. Saffell
Imperial College London, Division of Cell & Molecular Biology,
London, UK
Large classes and limitations in time and equipment mean that
university undergraduate bioscience laboratory practical classes
often involve students following recipes in the hope of achieving
a ‘correct’ result. This does not simulate the creative and investi-
gative nature of the research process. Moreover, it gives students
a misleading perception of the nature of bioscience research, and
what it is like to be a bioscience researcher. Students who have
the chance to experience genuine research in our labs are often
inspired to consider a bioscience research career, and feeling
included within the research community is cited by many as an
important consideration in their decision. To simulate the
research process for larger groups, I designed a third-year module
in which students plan, design and execute their own original
research in small groups within given parameters. Interactive
experimental design tutorials preceding the wet-lab phase take
students from the principles of good experimental design and sty-

lised problem scenarios through to iterative experimental design
and planning of their own experiments, including peer feedback
and comment. After the wet lab phase, tutorials are given on
data analysis and how to present scientific data in the form of a
paper. Evaluation indicates that the module is highly valued, and
it is the freedom to design their own experiments that students
value most. The talk will present the module rationale, design,
style and key elements, and highlight the perceptions of students
on its value to them.
I114
PI3KC2a, a class II PI3K, is required for
dynamin-independent internalization
pathways
A. E. Salcini
University of Copenhagen, BRIC, Copenhagen, Denmark
Endocytosis of molecules from the extracellular milieu can occur
through distinct internalization routes. Clathrin and caveolin-med-
iated pathways are by far the best characterized and they require
dynamin, a large GTPase responsible for the scission of newly
formed vesicles at the plasma membrane. One or more endocytic
mechanisms occurring independently of clathrin and caveolin have
been identified but are still depending on functional dynamin.
Abstracts Invited speakers
26 FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies
However, a growing number of molecules are reported to internal-
ize in a dynamin-independent manner, as is the case for GPI-
anchored proteins (GPI-APs), MHC1 and various bacterial toxins
and viruses, but the molecular players regulating dynamin-inde-
pendent endocytosis and the following trafficking steps are still
largely unknown. In a survival-based sh-RNA screen we identified

several molecules as putative regulators of dynamin-independent
pathways and PI3KC2a, a class II PI3K, was further validated.
Our results indicate that: (i) PI3KC2a colocalizes with cargo pro-
teins internalizing through dynamin-independent pathways and re-
localizes to endomembranes during dynamin-independent
internalization. (ii) PI3KC2a positively regulates physiological
internalization events occurring in a dynamin-independent fashion
such as fluid phase endocytosis and internalization of CD59. (iii)
PI3KC2a action is mediated by EEA1 recruitment in internalizing
vesicles, a step required for the internalization and intracellular
trafficking of vesicles generated by dynamin-independent pro-
cesses. The role of PI3KC2a and other identified proteins during
dynamin-independent internalization events will be discussed.
I115
Circadian clock control of the cellular response
to DNA damage
A. Sancar, L. Lindsey-Boltz, T-H. Kang, J. Reardon, J. H. Lee
and N. Ozturk
Department of Biochemistry and Biophysics, University of North
Carolina, Chapel Hill, NC, USA
Mammalian cells possess a cell-autonomous molecular clock
which controls the timing of many biochemical reactions and
hence the cellular response to environmental stimuli including
genotoxic stress. The clock consists of an autoregulatory tran-
scription-translation feedback loop made up of four genes/pro-
teins, BMal1, Clock, Cryptochrome, and Period. The circadian
clock has an intrinsic period of about 24 h, and it dictates the
rates of many biochemical reactions as a function of the time of
the day. Recently, it has become apparent that the circadian
clock plays an important role in determining the strengths of cel-

lular responses to DNA damage including repair, checkpoints,
and apoptosis. These new insights are expected to guide develop-
ment of novel mechanism-based chemotherapeutic regimens.
I116
Protein toxins from plants and bacteria:
probes for intracellular transport and tools in
medicine
K. Sandvig
Department of Biochemistry, Institute for Cancer Research, The
Norwegian Radium Hospital, Oslo University Hospital, Oslo,
Norway
A number of protein toxins produced by bacteria and found in
plants enter eukaryotic cells and inhibit protein synthesis enzymat-
ically. These toxins include the bacterial toxin Shiga toxin pro-
duced by Shigella dysenteriae, the related Shiga-like toxins
produced by E. coli, as well as the plant toxin ricin. Bacterial infec-
tions associated with toxin production are still a problem world-
wide. However, although being a threat to human health, toxins
are valuable tools to discover and characterize cellular processes
such as endocytosis and intracellular transport. Importantly, they
can also be exploited in cancer diagnosis and therapy. After bind-
ing to cell surface receptors, toxins are endocytosed by different
mechanisms (both clathrin-dependent and -independent routes),
and from endosomes they can be transported retrogradely to the
Golgi apparatus and then to the ER before translocation to the
cytosol. Sorting of the toxins between endosomes and the Golgi
apparatus are regulated by the membrane lipid composition as well
as protein complexes and kinases. The requirement for transport is
more strictly regulated for the glycosphingolipid (Gb3)-binding
toxin Shiga, than for ricin which binds to both lipids and proteins

with terminal galactose. The interaction between toxins and cells
and their possible use in medicine will be discussed.
I117
Transcriptional control of metabolic fluxes and
computational identification of the governing
principles
R. Kleijn, S-M. Fendt, R. Schuetz, M. Heinemann, N. Zamboni
and U. Sauer
ETH Zurich, Zurich, Switzerland
Great strides have been made in our ability to monitor transcrip-
tional and proteome responses, but it is not trivial to predict bio-
logical function and activity from such data [1]. Here we elucidate
which transcription factors actually control metabolic function in
a given environment on the basis of large-scale flux analyses [2]
upon genetic and environmental perturbations in E. coli and B.
subtilis. Systematic flux analysis of 120 transcription factor
mutants in the yeast S. cerevisiae then reveal networks of active
transcriptional regulation under five conditions. Using metabolo-
mics and targeted proteomics for in depth analysis of key mutants
then identify the precise molecular targets of these active regula-
tion mechanisms. While many proteins are differentially expressed
in deletions mutants and under different conditions, only rela-
tively few of these expression changes actually cause flux altera-
tions, as revealed by computational analyses of flux, proteome
and metabolome data. On the basis of these large in vivo flux
data, we then ask whether there are generally valid principles that
describe the distribution of flux under different conditions and
how such metabolic networks respond to perturbations? Using
the computational framework of flux balance analysis, we test
two fundamentally different families of hypotheses: are cells opti-

mized during evolution towards one or more objectives [3] or are
their responses optimized towards minimal readjustments?
References:
1. Heinemann M & Sauer U. Curr Opin Microbiol 2010.
2. Fischer E. & Sauer U. Nat Genet 2005; 37: 636–640.
3. Schu
¨
tz R, Ku
¨
pfer L & Sauer U. Mol Syst Biol 2007; 3:119.
I118
Biglycan signaling: Toll-like receptors, the
NLRP3-inflammasome and beyond
L. Schaefer
Institute of Pharmacology, University of Frankfurt, Frankfurt/
Main, Germany
The role of endogenous inducers of inflammation is poorly
understood. To produce the proinflammatory master cytokine
IL-1, macrophages need double stimulation with ligands to both
Toll-like receptors (TLR) for IL-1 gene transcription and nucleo-
tide-binding oligomerization domain-(NOD)-like receptors for
activation the inflammasome. It is particularly intriguing to
define how this complex regulation is mediated in the absence of
an infectious trigger. Biglycan, an ubiquitous leucine-rich repeat
(LRR) proteoglycan of the extracellular matrix, interacts with
TLR2/4 on macrophages. Objective of this study was to define
the role of biglycan in the synthesis and activation of IL-1. Here
we show that in macrophages soluble biglycan induces the
NLRP3/ASC inflammasome, activating caspase-1 and releasing
mature IL-1 without need for additional costimulatory factors.

Invited speakers Abstracts
FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies 27
This is brought about by the interaction of biglycan with TLR2/
4 and purinergic P2X4/P2X7 receptors, which induces receptor
cooperativity. By signaling through TLR2/4 biglycan stimulates
the expression of NLRP3 and pro-IL-1 mRNA. Both in a model
of non-infectious inflammatory renal injury (unilateral ureteral
obstruction) and in LPS-induced sepsis biglycan-deficient mice
displayed lower levels of active caspase-1 and mature IL-1 in the
kidney, lung and circulation. Our results provide evidence for
direct activation of the NLRP3 inflammasome by biglycan and
describe a fundamental paradigm of how tissue stress or injury is
monitored by innate immune receptors detecting the release of
the extracellular matrix components and turning such a signal
into a robust inflammatory response.
I119
Abstract missing, please view Abstract Addendum
I120
Separation of centrosomes during the cell
cycle
B. Mardin
1
, A. M. Fry
2
and E. Schiebel
1
1
Center for Molecular Biology (ZMBH), University of
Heidelberg, ZMBH, Heidelberg, Germany,
2

Department of
Biochemistry, University of Leicester, Leicester, UK
The centrosome can be envisioned as a molecular machine that
has important functions in microtubule organization, cell cycle
regulation, establishment of cell polarity and stem cell division. It
is also a platform for signal transduction molecules. One such
signal transduction network whose components are associated
with the centrosome is the Hippo pathway. The Hippo pathway
has a well-established function in tissue growth control and
apoptosis. However, the importance of Hippo components at
centrosomes is less clear. Here, we report that two Hippo path-
way components, the mammalian sterile 20-like kinase 2 (Mst2)
and the scaffold protein Salvador (hSav1), regulate the NIMA-
related kinase Nek2A in its ability to initiate centrosome disjunc-
tion. Nek2A has a SARAH-like C-terminal coiled-coil region
that mediates direct interactions with the SARAH domains of
hSav1 and Mst2. Mst2 and hSav1 are important for the centroso-
mal localization of Nek2A and for Nek2A-dependent phosphory-
lation of C-Nap1 and rootletin, the centrosomal linker proteins
whose phosphorylation triggers centrosome separation. Mst2
directly phosphorylates Nek2A and non-phosphorylated forms of
Nek2A fail to induce centrosome splitting, while the phospho-
mimicking Nek2A mutants are able to split centrosomes even in
the absence of Mst1/2 activity. We propose that hSav1 and Mst2
regulate localized activity of Nek2A at centrosomes in early mito-
sis in response to cell cycle signals.
I121
Cell cycle control during growth and
reproduction
A. Schnittger

Institut de Biologie Mole
´
culaire des Plantes du CNRS, Molecular
Mechanisms of Plant Plasticity, Strasbourg, France
Proper control of cell proliferation is of key importance for any
multicellular organism and requires a tight coordination with
growth and differentiation. At the heart of the eukaryotic cell
cycle are cyclin-dependent kinases (CDKs) that regulate the entry
into the next cell-cycle phase. In particular, high levels of CDK
activity are required for entry and progression of DNA synthesis
phase (S-phase) and for execution of mitosis (M-phase). CDKs
function as microprocessors and integrate extrinsic cues, such as
nutrient availability, with intrinsic cues, for instance the develop-
mental program and only if a certain threshold of CDK activity is
reached will the entry into the next cell-cycle phase be promoted.
Here, I will focus on the control and function of the major cell
cycle regulator CDKA;1, the Arabidopsis homolog of the yeast
Cdc2/CDC28 kinase. CDKA;1 is required throughout plant devel-
opment, starting from meiosis to the development of the gameto-
phytes, embryonic and postembryonic growth. Comparing the
action and regulation of CDKs in plants with what has been
found in other model organisms shows that the mechanistics of
Cdc2-like kinases are conserved. However, the regulatory context
and the wiring appear to be species-specific adapted.
I122
Predicting pathways in genome-scale
metabolic networks
S. Schuster, L. F. de Figueiredo and C. Kaleta
Department of Bioinformatics, University of Jena, Jena,
Germany

Metabolic pathway analysis has become an important part of
Systems Biology. The concept of elementary modes is a very use-
ful tool in that analysis. It allows one to decompose complex
metabolic networks into the smallest functional entities. This has
manifold applications in bioengineering, such as microbial strain
improvement and determining the robustness against knockouts.
We discuss an example of a successful theoretical prediction of a
novel pathway in central metabolism in Escherichia coli, the
PEP-glyoxylate cycle. Pathway analysis faces the problem of the
combinatorial explosion of the number of elementary modes with
increasing system size, which has hampered scaling it up to gen-
ome-wide metabolic models. We present two novel approaches to
overcoming this obstacle. One approach is by computing first the
shortest elementary mode, then the second-shortest etc., up to a
certain length. This can be done by integer linear optimization.
The second concept is called elementary flux patterns. Within a
large metabolic network, elementary flux patterns are defined as
sets of reactions that represent the basic routes of any steady-
state flux of the network through a particular subsystem that are
compatible with admissible fluxes in a (possibly) much larger
metabolic network. The subsystem can be made up by reactions
in which we are interested in, for example, reactions producing a
certain metabolite. This allows one to predict metabolic pathways
in genome-scale networks. We will present several examples such
as the frequently neglected glycolate pathway and the role of the
GABA shunt as a bypass of part of the TCA cycle in E. coli,
which has been known before from plants.
I123
The endocytic networks in the control of the
plasticity of tumor cell migration

G. Scita
Ifom Foundation, Milan, Italy
Tumor cells can adopt different modes of cell motility. The abil-
ity to switch between diverse modes of migration enables tumors
to adapt to micro-environmental conditions and to metastasize.
The critical pathways and cellular processes underlying the plas-
ticity of tumor cell motility have only begun to be identified. An
appealing hypothesis, supported by recent evidence, is that endo-
cytosis, originally thought of as a device to internalize nutrients
and membrane-bound molecules, is a connectivity infrastructure
(which we call ‘the Endocytic Matrix’) of different cellular net-
works necessary for the execution of various cellular programs.
Abstracts Invited speakers
28 FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies
A primary role of the Endocytic Matrix is the delivery of space-
and time-resolved signals to the cell, and it is thus essential for
the execution of polarized functions during 3D cell migration
and invasion. Here, by focusing on the endocytic and signaling
functions of Rab5, a small GTPases essential for endosome bio-
genesis, we will discuss recent experimental evidence that support
the general paradigm that intracellular trafficking, controlled by
Rab5, is needed to re-direct molecules to restricted regions of the
plasma membrane, enabling sustained and polarized signalling
for directed cell migration and invasion, ultimately regulating the
plasticity of tumor cell motilty.
I124
Size matters: mitochondria elongate to
support survival of starving cells
L. Gomes
1

, G. Di Benedetto
2
and L. Scorrano
3
1
Dulbecco Telethon Institute, Venetian Institute of Molecular
Medicine, Padova, Italy,
2
Venetian Institute of Molecular
Medicine, Padova, Italy,
3
University of Geneva, Cell Physiology
and Metabolism, Geneva, Switzerland
The multiple roles of mitochondria in cell metabolism, prolifera-
tion and death are matched by their complex and dynamic mor-
phology. Changes in mitochondrial shape and ultrastructure are
hallmarks of, and required for, apoptosis. During nutrient restric-
tion, autophagy sustains the energetic demand of the cell, by
allowing the recycling of its components. It is unclear whether
autophagy is accompanied by changes in mitochondrial morphol-
ogy. Here we show that mitochondrial elongation supports effi-
cient energy production and survival during autophagy.
Unexpectedly, induction of autophagy led to mitochondrial elon-
gation both in vitro and in vivo. Elongation correlated with
increased fusion rate and required the core mitochondrial fusion
proteins. Starvation triggered a rise in cellular cAMP levels and
the activation of protein kinase A (PKA), which blocked translo-
cation of the pro-fission protein DRP1 to mitochondria. During
starvation,elongated mitochondria were spared from mitophagy,
possessed more cristae, and were more efficient in ATP produc-

tion, as a result of increased dimerization and activity of the
ATP syntase. Mitochondria unable to elongate were conversely
dysfunctional and consumed ATP, leading to starvation-induced
death. Our data indicate that elongated mitochondria are
required for ATP maintenance and survival of starving cells.
I125
Abstract missing, please view Abstract Addendum
I126
Research possibilities for pre-graduate
students at the Faculty of Medicine, University
of Lisbon: over 10 years of experience with the
GAPIC programme
A. Sebastia
˜
o
1
and A. Barroso
2
1
Faculty of Medicine, and Unit of Neurosciences, Institute of
Molecular Medicine, Institute of Pharmacology and Neurosciences,
University of Lisbon, Lisboa, Portugal,
2
GAPIC, Faculty of Medi-
cine, University of Lisbon, Lisboa, Portugal
The Faculty of Medicine at the University of Lisbon started, in
1996, a programme aiming to allow the direct and active contact
of medical students with scientific research. During 1 year, with a
realistic schedule defined by the student and the supervisor, the
student develops a small project with a well defined objective and

research question. At the end of the year each student has to
present the results at a workshop at the Medical School. A writ-
ten scientific report is also asked. This activity is optional and no
other school duties are waived. A student can join the pro-
gramme just for 1 year or can re-apply for it. Some students pre-
fer to follow a research track in successive years; some others
prefer to experience different research topics or perspectives (eg
basic research vs clinical research). Projects rated as good, very
good or excellent by an evaluation panel are usually accepted
and supported (maximum 2500€ per project, given to the
research lab). At 2009/2010, there are 40 projects running. Maxi-
mum number of students involved at each project is 3, but most
projects have only one student. A follow up survey on the impact
of the programme in the years following graduation showed that
about 1/3 of the students engaged in the programme during any
year during pre-graduation are enrolled in research activities after
leaving the Medical School.
I127
Differential innate immune signalling via Ca
2+
sensor protein kinases in plants
M. Boudsocq, M. Willmann, M. McCormack, H. Lee, L. Shan,
P. He, J. Bush, S-H. Cheng and J. Sheen
Massachusetts General Hospital, Boston, MA, USA
Innate immunity represents the first line of inducible defense
against microbial infection in plants and animals. In both king-
doms, recognition of pathogen- or microbe-associated molecular
patterns (PAMPs or MAMPs), such as flagellin, initiates conver-
gent signalling pathways involving MAP kinase (MAPK) cas-
cades and global transcriptional changes to boost immunity.

Although calcium has long been recognized as an essential and
conserved primary mediator in plant defense responses, how cal-
cium signals are sensed and relayed into early MAMP signalling
is unknown. We use a functional genomic screen and genome-
wide gene expression profiling to show that four calcium-depen-
dent protein kinases (CDPKs) are calcium sensor PKs critical to
transcriptional reprogramming in plant innate immune signalling.
Unexpectedly, CDPKs and MAPK cascades act differentially in
four MAMP-mediated regulatory programs to control early
genes involved in synthesis of defense peptides and metabolites,
cell wall modifications and redox signalling. Transcriptome pro-
file comparison suggests that CDPKs are the convergence point
of signalling triggered by most MAMPs. Double, triple and qua-
druple cpk mutant plants display progressively diminished oxida-
tive burst and gene activation induced by flg22, as well as
compromised pathogen defense. In contrast to negative roles of
calmodulin (CAM) and a CAM-activated transcription factor in
plant defense, the present study reveals calcium signalling com-
plexity and demonstrates key positive roles of specific CDPKs in
initial MAMP signalling.
I128
Fighting disease by selective autophagy of
aggregate-prone proteins
A. Simonsen
Institute of Basic Medical Sciences, University of Oslo, Oslo,
Norway
Intracellular aggregation of ubiquitinated proteins is a major cyto-
pathological feature of most neurodegenerative disorders, includ-
ing Alzheimer’s disease, Parkinson’s disease and Huntington’s
disease. Over the past several years, it has become increasingly evi-

dent that macroautophagy drives the elimination of accumulated
protein even in the absence of nutrient-deprivation. It has yet to
be determined, however, whether the elimination of these proteins
Invited speakers Abstracts
FEBS Journal 277 (Suppl. 1) 5–36 (2010) ª 2010 The Authors Journal compilation ª 2010 Federation of European Biochemical Societies 29