Chapter 13: Gene Expression
13
Gene Expression
I.
Making proteins
A. Garrod (1908) proposed that genes are responsible for enzymes
1.
Studying a rare genetic disease, alkaptonuria
2.
Symptom—homogentisic acid accumulate in urine turning it black with air
3.
Autosomal recessive
4.
Missing enzyme necessary to break tyrosine  CO2 + H2O
A. Beadle and Tatum (1940)—fig 13-2
II.
1.
Neurospora (bread mold)
2.
Grows mostly haploid
3.
Expose strains to x-ray and UV
4.
Identified mutant strains
5.
One-gene—one enzyme
6.
revised to one gene—one polypeptide hypothesis
Overview Protein Synthesis
A. Transcription
1.
DNA is transcribed to form mRNA
2.
RNA is a single-stranded, uncoiled nucleotide polymer
3.
The sugar in RNA is ribose, and uracil substitutes for thymine
a)
Uracil is a pyrimidine and forms two bonds with adenine
13 - 1
B.
Translation
1.
involves mRNA, tRNA (transfer RNA), and rRNA (ribosomal RNA)
2.
mRNA has sequences of 3 nucleotides called codons (fig 13-5)
a)
triplet code-- codons are read in sequences of 3
b)
Codons are written in a 5' to 3' fashion
c)
Each codon codes for 1 amino acid
d)
4 bases can combine in 43 combinations—more than enough to code
for the 20 naturally occurring amino acids – 64 words
3.
Each tRNA molecule has a sequence of 3 nucleotides—the anticodon
a)
Anticodons base pair with the codon in a complimentary way
b)
Anticodons are written in a 3' to 5' fashion
4.
Ribosomes are composed of proteins and rRNA
5.
The genetic code is redundant
6.
a)
Redundant codons have similar base sequences
b)
Only methionine and tryptophan have single triplet codes
The wobble hypothesis-- the third nucleotide of the tRNA anticodon can
bind with more than one nucleotide of the mRNA codon, as cells typically
produce only about 40 different tRNA molecules
III.
Transcription fig 13-8 ****
A. RNA is synthesized by DNA-dependent RNA polymerases
B.
Require an input of ATP
13 - 2
Chapter 13: Gene Expression
C.
Messenger RNA contains base sequences that code for protein
1.
Transcription does NOT involve a primer
2.
Begins at a promoter sequence on the template strand (the promoter is
NOT transcribed)
3.
RNA synthesis proceeds in a 5'3' direction (copying the DNA from the 3'
 5')
a)
Upstream
b)
Downstream
D. Steps in synthesis of mRNA
1.
Initiation
a)
RNA polymerases move past the promoter to begin transcription of
DNA
b)
Bacterial promoters are about 40 bases long and are located in the
DNA, just upstream from the starting point
c)
2.
3.
Unlike DNA synthesis, RNA synthesis does not require a primer
Elongation
a)
nucleotides are added with the input of energy
b)
continues until RNA polymerases recognize a stop signal
Termination
a)
At the stop signal, the process differs in prokaryotes and eukaryotes
b)
In prokaryotes, transcription stops immediately
13 - 3
c)
In eukaryotes, RNA polymerase continues to add new nucleotides for
about 10–15 nucleotides past the stop
4.
Typically only one strand of the DNA is transcribed—the template strand
a)
E.
some of the template sections are on one strand or the other strand
Messenger RNA contains base sequences that do NOT directly code for
protein
1.
Noncoding leader sequence at the 5' end, important in binding to the
ribosome during translation
IV.
2.
The start codon (AUG) begins the coding sequences
3.
Stop codon (UAA, UAG, or UGA), and
4.
Eukaryotes add variable noncoding 3' trailing sequences
Translation fig 13-13,14,15
A. An amino acid is attached to tRNA
B.
Sequence of amino acids determined by sequence of codons on mRNA
C.
tRNA
1.
Attachment sites for amino acids
2.
Produced from tRNA genes
a)
tRNA is linked to amino acids by aminoacyl-tRNA synthetases,
forming aminoacyl-tRNAs
b)
This is an energy-requiring process
13 - 4
Chapter 13: Gene Expression
3.
Bind to complementary codons of the mRNA
4.
Must be recognized by both the specific aminoacyl-tRNA synthetase and
the ribosome and have recognition sites for each
5.
tRNA molecules are approximately 70 nucleotides long, with some generic
sections and some unique sections
6.
The nucleotide chain is folded back upon itself to form 3 or more loops,
with unpaired nucleotides exposed to form the recognition or attachment
sites
D. The components of the translational machinery come together at the
ribosomes
1.
Ribosomes are composed of two subunits
a)
The large subunit has a groove into which the small subunit fits
b)
Ribosomes are transcribed from DNA but do not carry information—
function as the physical site of translation and as a catalyst
c)
The A (aminoacyl) site of the ribosome is where the aminoacyl-tRNA
binds
d)
The P (peptidyl) site of the ribosome is where the tRNA holding the
polypeptide chain is positioned
e)
The E (exit) site is where the tRNAs that have delivered the amino
acids to the growing polypeptide chain exit the ribosome
E.
Translation process ****
13 - 5
1.
Initiation
a)
Initiation factors (proteins) move an initiation tRNA onto the small
ribosomal subunit
b)
The codon for the initiation is AUG, which codes for the amino acid
methionine (met), met is often removed later
c)
The initiation complex binds to ribosome-recognition sequences on
the mRNA and aligns the anticodon of the tRNA with the codon of
the mRNA
d)
The large ribosomal subunit then binds, forming the functional
ribosome or initiation complex
e)
In eukaryotes, the met of the initiator tRNA is unmodified; there is no
leader sequence, but a start codon is embedded in a short sequence;
and lastly, the initiation complex is more complex
2.
Elongation
a)
tRNA is bound to the P site of the ribosome; the A site is unoccupied
until the next aminoacyl-tRNA moves in
b)
The energy for this binding process (GTP)
c)
The tRNA molecule is released from the P site
(1) This reaction requires the ribozyme, peptidyl transferase
d)
Protein synthesis always proceeds from the amino end to the carboxyl
end
e)
Translocation
13 - 6
Chapter 13: Gene Expression
F.
f)
tRNA, without an attached amino acid, moves from the P to the E site
g)
tRNA then leaves the ribosome
h)
tRNA is reused
Termination
1.
mRNA presents the codons UAA, UGA, or UAG
2.
Release factors recognize codons, breaking bond between the tRNA in the
P site
3.
G.
V.
Ribosome dissociates into 2 units
Polyribosome is a complex of one mRNA and many ribosomes
1.
In prokaryotes, transcription and translation occur simultaneously
2.
The half-life of the mRNA molecules in bacteria is about 2 minutes
3.
half-lives of eukaryotic mRNA molecules are as long as 24 hours
Transcription and translation are more complex in eukaryotes
A. Eukaryotic mRNA molecules are processed after transcription; prokaryotic
mRNAs are used immediately without processing
1.
Processing begins in the nucleus; 5' cap is added to the 5' end
a)
The cap is 7-methylguanylate, which is GMP and a methyl group
added
b)
2.
The cap is necessary for binding to ribosomes
Capping may protect mRNA from degradation;
13 - 7
3.
At the 3' end of the molecule, a sequence is added that acts as a signal for
the subsequent addition of a tail
4.
B.
Enzymes recognize the signal and add 100–250 adenines (the poly-A tail)
Noncoding sequences and coding sequences transcribed from eukaryotic
genes
1.
C.
Genes of most eukaryotes have interrupted coding sequences fig13-17
a)
Exons (expressed sequences)
b)
Introns (intervening sequences)
2.
Precursor mRNA (pre-mRNA) contains both introns and exons
3.
Small nuclear ribonucleoprotein complexes (snRNPs)
4.
RNA may also act as a self-acting enzyme and excise and splice
Revised definition of a gene
1.
A gene is a nucleotide sequence which produces a specific RNA or a
polypeptide
D. Reverse transcriptase
VI.
1.
Retro-Viruses
2.
HIV-1
3.
tool for molecular biologists
Mutations
A. Base-substitution
1.
may not alter the product
13 - 8
Chapter 13: Gene Expression
2.
3.
B.
C.
Missense mutations
a)
Nonfunctional
b)
Silent mutation
Nonsense mutations result in a stop codon
Frameshift mutations
1.
change the reading frame
2.
result in a nonfunctional protein
Transposons, or transposable genetic elements
1.
may disrupt genes but may also activate some genes
D. Mutagens
13 - 9