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Chapter 17—From Gene to
Protein
“Let me out of the nucleus, finally!”
or
The functional definition of a gene
I. The Connection Between Genes & Proteins
Metabolic defects = non-functional enzymes
• Studying metabolic diseases suggested that
genes specified proteins
- alkaptonuria (black urine from alkapton)
- PKU (phenylketonuria)
• Conclusion: Genes dictate phenotype
Beadle and Tatum (1941)
Experiments with Neurospora
ARGININE MUTANTS
One Gene-One Enzyme Hypothesis (Beadle & Tatum)
The function of a gene is to dictate the production of a specific enzyme
One Gene—One Enzyme� but not all proteins are enzymes� those proteins are coded by genes too
One Gene—One Protein�but many proteins are composed of several polypeptides, each of whichhas its own gene
One Gene—One Polypeptide
CENTRAL DOGMA of Molecular Biology
How does the information get from nucleus to cytoplasm?
Messenger RNA (mRNA)
= bridge between DNA & protein
DNA vs. RNA
RNA
DNA
StrandedNucleotidesSugar
Transcription—synthesis of RNA
under the direction of DNA• 1 DNA strand is template strand
• complementary RNA strand
is made– messenger RNA (mRNA)
• Enzyme = RNA polymerase
Translation:• Synthesis of a polypeptide under the
direction of mRNA
DNA → RNA → Protein
Prokaryotes vs. Eukaryotes
The Genetic Code
3 letter codon =
genetic “word”
Each word stands
for an amino acid
AUG = start codon
(starts a protein,
adds a methionine)
UAA/UAG/UGA =
stop codons
(signals end of
protein)
Code is universal—
from bacteria to
plants & animals (evolved very early
in the history of life)“Redundancy without Ambiguity”
II. The Synthesis & Processing of RNA
Transcription
Step 1 Initiation:RNA polymerase binds to promotersequence on DNA
Step 2 Elongation:RNA is made 5´→ 3´
Step 3 Termination:RNA polymerase stops at terminatorsequence
mRNA leaves nucleus through pores after processing
Transcription Animation
http://www-class.unl.edu/biochem/gp2/m_biology/animation/gene/gene_a2.html
Initiation of Transcription at a Eukaryotic Promoter
Transcription factors bind to
DNA
THEN…
RNA Polymerase binds
Transcription Factors =proteins which bind to DNA & turn transcription “on” or “off” (master regulators)
Transcription Unit = length of DNA that is transcribed into RNA
RNA Processing (only in eukaryotes!)
Step 1: Alteration of mRNA ends-- add 5’ cap (modified G (GTP))
-- protects RNA from degradation (hydrolytic enzymes in cytoplasm)-- “attach here” sign for ribosomes
-- add polyA tail (50-250 A’s)-- same as above ☺☺☺☺ and…-- helps mRNA export from nucleus
Step 2: RNA Splicing (cut & paste)-- Pre-mRNA → mRNA
-- edit out introns (noncoding regions)-- splice together exons (coding sequences)
In higher eukaryotes 90% or more of gene can be intronno one knows why…yet…
Spliceosome = snRNPs & proteins
� Recognize splice sites
� Ribozymes catalyze splicing process
Exon = protein domain
Alternative RNA splicing…
III. Synthesis of Protein
How are the codons read?
Translation Basics
� Ribosome reads mRNA in codons
� tRNA anticodon base pairs with codon of mRNA
� tRNA brings in correct amino acid
� Amino acids assembled into polypeptide chain
Structure of Transfer RNA (tRNA)
“clover leaf” structure
�anticodon on “clover leaf”end
�amino acid on 3’ end
Aminoacyl-tRNA Synthetases
Enzyme that bonds amino acids to tRNAs
Ribosomes
Consist of proteins and ribosomal RNA (rRNA)
2 subunits (large & small)
Made in nucleolus (eukaryotes) and then exported
Figure 17.15 pg. 316
P site (peptidyl-tRNA site)holds tRNA carrying growing polypeptide chain
A site (aminoacyl-tRNA site)holds tRNA carrying next amino acid to be added to chain
E site (exit site)discharged tRNA leaves ribosome from exit site
TRANSLATION DETAILSStep 1: Initiation—requires small ribosomal subunit, Met tRNA, large ribosomal subunit, initiation factors (proteins), & GTP (energy)
Step 2:
Elongation
Amino acids are added one by one to the preceding A.A.
Occurs in a 3-step cycle with the help of elongation factors (proteins):
1. Codonrecognition(A site)
2. Peptide bond formation
3. Translocation (A to P site)
Ribosome moves 5’ → 3’on mRNA
Step 3: Termination
stop codon in mRNA reaches A site
release factor (protein) binds to stop codon
protein is freed from the ribosome
other components disassemble
Polyribosomes—
Clusters of ribosomes
that translate a single mRNA simultaneously
Many copies of a protein are made
quickly
(prokaryotes &
eukaryotes)
Coupled Transcription & Translation
Prokaryotes—
With no nucleus bacteria can simultaneously transcribe & translate the same gene
Posttranslational Modifications—(Polypeptide → Functional Protein)
chemically modified amino acids, removal of amino acids, polypeptide cut into pieces, or several joined together
Signal peptide—(postal code)stretch of amino acids that targets the protein to a specific destination in a eukaryotic cell
Point mutations can affect protein
structure and function
Mutations—changes in the genetic material of a cell
Point mutation–a change in 1 base pair of a gene
Example—sickle cell anemia
2 Categories of point mutations:
1. Substitutions
silent mutations—
same A.A. inserted
missense mutations—
different A.A. inserted
nonsense mutations—
Stop codon formed
2. Insertions/Deletions
frameshift mutation—
changes reading frame
causes shortened,
nonfunctional proteins
Mutagens?
Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
1. A mutation results in a defective enzyme a. In the following simple metabolic pathway, what would be
a consequence of that mutation?
a) an accumulation of A and no production of B and C
b) an accumulation of A and B and no production of C
c) an accumulation of B and no production of A and C
d) an accumulation of B and C and no production of A
e) an accumulation of C and no production of A and B
ca b
enzyme benzyme a
Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
3. A portion of the genetic code is UUU = phenylalanine, GCC = alanine, AAA = lysine, and CCC = proline. Assume
the correct code places the amino acids phenylalanine,
alanine, and lysine in a protein (in that order). Which of the following DNA sequences would substitute proline for
alanine?
a) AAA-CGG-TTA
b) AAT-CGG-TTT
c) AAA-CCG-TTT
d) AAA-GGG-TTT
e) AAA-CCC-TTT
Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
4.A particular triplet of bases in the coding sequence
of DNA is AAA. The anticodon on the tRNA that
binds the mRNA codon is
a) TTT.
b) UUA.
c) UUU.
d) AAA.
e) either UAA or TAA, depending on wobble in the first base.
Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
5. A part of an mRNA molecule with the following sequence is
being read by a ribosome: 5' CCG-ACG 3' (mRNA). The
following activated transfer RNA molecules are available. Two of them can correctly match the mRNA so that a dipeptide can
form.
The dipeptide that will
form will be
a) cysteine-alanine.
b) proline-threonine.
c) glycine-cysteine.
d) alanine-alanine.
e) threonine-glycine.
CysteineACG
AlanineCGG
GlycineCCG
ThreonineUGC
AlanineCGU
ProlineGGC
Amino AcidtRNA Anticodon
Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
7. Each of the following is a modification of the sentence THECATATETHERAT.
A. THERATATETHECATB. THETACATETHERATC. THECATARETHERATD. THECATATTHERATE. CATATETHERAT
Which of the above is analogous to a frameshift mutation?
a) A
b) B
c) C
d) D
e) E
Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
8. Each of the following is a modification of the sentence THECATATETHERAT.
A. THERATATETHECATB. THETACATETHERATC. THECATARETHERATD. THECATATTHERATE. CATATETHERAT
Which of the above is analogous to a single substitution mutation?
a) A
b) B
c) C
d) D
e) E
Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
9. What is the relationship among DNA, a gene, and a chromosome?
a) A chromosome contains hundreds of genes, which are
composed of protein.
b) A chromosome contains hundreds of genes, which are
composed of DNA.
c) A gene contains hundreds of chromosomes, which are
composed of protein.
d) A gene is composed of DNA, but there is no relationship
to a chromosome.
e) A gene contains hundreds of chromosomes, which are
composed of DNA.
Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
10.A biologist inserts a gene from a human liver cell into
the chromosome of a bacterium. The bacterium then transcribes this gene into mRNA and translates the mRNA into protein. The
protein produced is useless.
The biologist extracts the protein and mature mRNA that codes
for it. When analyzed you would expect which of the following
results?
a) the protein and the mature mRNA are longer than in human
cells
b) the protein and mature mRNA are shorter than expected
c) the protein is longer and the mRNA is shorter than expected
d) the protein is shorter and the mRNA is longer than expected