GENE EXPRESSION THE FLOW OF GENETIC INFORMATION

6TH WEEK, BIO-1053

GENE EXPRESSION
THE FLOW OF GENETIC INFORMATION

6th week

General Genetics-BIO1053


Chapter outline
The genetic code
- How triplets of the four nucleotides unambiguously specify
20 amino acids, making it possible to translate information
from a nucleotide chain to a sequence of amino acids
Transcription: From DNA to RNA
Translation: From mRNA to Protein
Differences in Gene expression between prokaryotes and
eukaryotes
The effect of mutation on gene expression and gene function
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General Genetics-BIO1053


The genetic code
The four nucleotides encode 20 amino acids
By deduction:
-If only one nucleotide represented an amino acid: only for 4
amino acids
-If 2 nucleotides represented each amino acid: 42=16 possible

combination of couplets (16 amino acids)
-If 3 nucleotides represented each amino acid: 43=64 possible
combination of triplets, more than enough
Must be at least triplet combination that encode for amino
acids
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General Genetics-BIO1053


The genetic code

Triplet codons of
nucleotides represent
individual amino acids
61 codons represent
the 20 amino acids, 3
codons signify stop

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General Genetics-BIO1053


Evidence that a codon is composed of more than one
nucleotide, Yanofsky, 1960s

Different point mutations may affect the same amino acid
• Codons must contain >1 nucleotide
Each point mutation affects only one amino acid

• Each nucleotide is part of only one codon


Evidence that a codon is composed of more than one
nucleotide, Yanofsky, 1960s


Studies of frameshift mutations showed that codons
consist of three nucleotides

F. Crick and S. Brenner (1955)
Proflavin-induced mutations in
bacteriophage T4 rIIB gene
• Intercalates into DNA
• Causes insertions and
deletions
2nd treatment with proflavin can
create a 2nd mutation that
restores wild-type function
(revertant)
• Intragenic suppression


Different sets of T4 rIIB mutations generate either a mutant
or a normal phenotype

Codons must be read in
order from a fixed
starting point
Starting point establishes a

reading frame
Intragenic supression
occurs only when wild-type
reading frame is restored


Codons consist of three nucleotides read in a defined
reading frame


Cracking the code: Discovery of mRNA
1950s, studies in eukaryotic cells
Evidence that protein synthesis takes place in cytoplasm

•Deduced from radioactive tagging of amino acids
•Implies that there must be a molecular intermediate
between genes in the nucleus and protein synthesis in the
cytoplasm
Discovery of messenger RNAs (mRNAs), molecules for
transporting genetic information

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General Genetics-BIO1053


Synthetic mRNAs and in vitro translation determines
which codons designate which amino acids

1961-Marshall Nirenberg and

Heinrich Matthaei created
mRNAs and translated to
polypeptides in vitro
Polymononucleotides
Later, Har Gobind Khorana,
made polydinucleotides and
polytrinucleotides,
polytretranucleotides
-> synthesis of polypeptides


Cracking the genetic code with mini-mRNAs

Nirenberg and Leder
(1965)
Resolved
ambiguities in genetic
code
In vitro translation
with trinucleotide
synthetic mRNAs and
tRNAs charged with a
radioactive amino acid


Correlation of polarities in DNA, mRNA, and polypeptide
mARN có chiều 5’-> 3’ tương ứng với chiều đầu N-> C
của chuỗi polypheptide
Một sợi ADN khuôn
Một sợi còn lại là trình tự giống ARN

Các condon vô nghĩa gây sự kết thúc chuỗi
polypeptides, bao gồm - UAA, UAG, UGA


The genetic code: summary
The code consists of a triplet codons, each of which specifies
an amino acid
Codons are nonoverlapping
The code includes 3 stop codon: UAA, UGA, UAG that
terminate translation
The code is degenerate- more than one codon specifies the
same amino acid
Fixed starting point establishes a reading frame. AUG is
initiation codon


The genetic code: summary
5’-3’ direction of mRNA corresponds with N-terminus to Cterminus of polypeptide
Mutation modify message encoded in sequence
- Frameshift mutations change reading frame
- Missense mutations change codon of amino acid to another
one
- Nonsense mutations change a codon for an amino acids to a
stop codon


Do living cells construct polypeptides according to same
rules as in vitro experiment?
Yanofsky: Single-base substitutions can explain the altered
amino acids in trp− and trp+ revertants

Missense mutations
are single nucleotide
substitutions and
conform to the code


Proflavin treatment generates trp- mutants
Further treatment generatees trp+ revertants
Single base deletion (trp-) and an insertion causes reversion
(trp+)


The genetic code is almost, but not quite, universal
All living organisms use same basic genetics code
- Translational systems can use mRNA from another
organism to generate protein
- Comparisons of DNA and protein sequence reveal perfect
correspondence between codons and amino acids among
all organisms
Exceptional genetic codes found in ciliates and
mitochondria, for example: UAA, UAG code for glutamine
instead of stop codon; CUA specifies threonine instead of
leucine


Transcription: From DNA to RNA
RNA polymerase catalyzes transcription
Promoters are DNA sequences that provide the signal to
RNA polymerase for starting transcription

RNA polymerase adds nucleotides in 5’-to-3’ direction
• Formation of phosphodiester bonds using ribonucleotide
triphosphates (ATP, CTP, GTP, and UTP)
• Hydrolysis of bonds in NTPs provides energy for
transcription
Terminators are RNA sequences that provide the signal to
RNA polymerase for stopping transcription

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Initiation: The beginning of transcription
RNA polymerase binds to promoter sequence located near beginning
of gene
• Sigma (s) factor binds to RNA polymerase ( holoenzyme)
• Region of DNA is unwound to form open promoter complex
• Phosphodiester bonds formed between first two nucleotides

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Elongation: an RNA copy of the gen
σ factor separates from RNA polymerase ( core enzyme)
Core RNA polymerase loses affinity for promoter, moves in 3’-to-5’
direction on template strand
Within transcription bubble, NTPs added to 3’ end of nascent mRNA

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General Genetics-BIO1053


Termination: the end of transcription
Terminators are RNA sequences that signal the end of
transcription
• Two kinds of terminators in bacteria: extrinsic (require rho factor) and
intrinsic (don’t require additional factors)
• Usually form hairpin loops (intramolecular H-bonding)

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Information flow

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The promoters of 10 different bacterial genes
Most promoters are upstream to the transcription start point
RNA polymerase makes strong contacts at -10 and -35

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General Genetics-BIO1053


In eukaryotes, RNA is processed after transcription
Capping enzyme adds a "backward" G to the 1st nucleotide of a primary
transcript
Methyl transferase add methyl group to this G and to one or two of the
nucleotides

Transcribed
bases

6th week

General Genetics-BIO1053