Download Chapter 15: Translation of mRNA

Chapter 15: Translation of mRNA
Student Learning Objectives
Upon completion of this chapter you should be able to:
1. Understand the role of translation in gene expression.
2. Understand the genetic basis for protein synthesis and the key experiments which led
to the discovery of gene function.
3. Know the features of the genetic code.
4. Know the structure and function of proteins.
5. Understand the role of tRNA and ribosomes in translation.
6. Understand the initiation, elongation, and termination stages of translation.
7. Distinguish between translation in bacteria and eukaryotes.
15.1 The Genetic Basis for Protein Synthesis
Overview
The final stage in the information flow from cellular genome to phenotype is the
manufacturing of a functional protein. Before going into the details of translation, it is important
to get a historical perspective, and this first section does just that. The section considers early
experiments which showed that the role of some genes is to encode enzymes.
In the early twentieth century, Archibald Garrod, a British physician, studied the
inherited disease alkaptonuria (Refer to Figure 15.1). Garrod described the disease as an inborn
error of metabolism. Indeed, he was the first to suggest that a connection exists between the
function of genes and the production of enzymes. The section then discusses the work of George
Beadle and Edward Tatum. In the early 1940s, they studied Neurospora mutants that were altered
in their nutritional requirements. A series of experiments such as the one shown in Figure 15.2,
allowed them to propose the one-gene/one-enzyme hypothesis. In later decades, this hypothesis
had to be modified in three ways: 1) some proteins are not enzymes; 2) some proteins are
composed of two or more different polypeptides; and 3) some genes encode RNAs that are not
translated into proteins (refer back to Table 14.1).
Outline of Key Terms
One gene-One enzyme hypothesis
Protein
Polypeptide
Translation
Structural gene
Messenger RNA (mRNA)
Alkaptonuria
Inborn error of metabolism
Focal Points


The metabolic pathway of phenylalanine breakdown (Figure 15.1)
An example of an experiment that supported Beadle and Tatum’s one-gene/one-enzyme
hypothesis (Figure 15.2)
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Exercises and Problems
Complete the following sentences with the most appropriate word or phrase:
(1) ________ was the first researcher to suggest that a relationship exists between the
function of a gene and the production of an enzyme. He studied the disease (2) ________, which
he described as an inborn error of metabolism. In this disorder, the patient’s body accumulates
abnormal levels of (3) _______ acid, which is excreted in urine.
The one-gene one-enzyme hypothesis was introduced in the 1940s by (4) _________.
They studied the inheritance of defects in the metabolic pathways of (5) ________,
The term (6) _______ refers to a structure; it is a linear sequence of amino acids. By
comparison, the term (7) _______ denotes function.
15.2 The Relationship Between the Genetic Code and Protein Synthesis
Overview
This section begins by exploring the relationship between transcription and protein
synthesis (Figure 15.3). You should use this figure as a reference for the remainder of the chapter.
In order for the genetic information, which is written as nucleic acids, to be converted to a
functional protein, a code must be present so that groups of nucleotides dictate specific amino
acids. This code is presented in Table 15.1. The section concludes with a discussion of amino
acid structure and the linking of amino acids to form three-dimensional proteins. One important
concept is the directionality of the mRNA to the structure of the polypeptide. Notice that the 5′
end of the mRNA corresponds to the amino (N-terminal) end of the polypeptide, while the 3′ end
of the mRNA is associated with the C-terminal end (Figure 15.4). You also need to become
familiar with the four levels of protein structure (Figure 15.7). This figure is especially important
since variations in the genetic information may produce alterations in the protein structure. This
may change the phenotype of the organism, alter a metabolic pathway, or in some cases cause
lethality. Thus, you need to firmly understand that the DNA and functional proteins are linked.
Outline of Key Terms
Amino acid
Side chain (R group)
Peptide bond
N-terminus (Amino-terminal end)
C-terminus (Carboxyl-terminal end)
Primary structure
Amino acid sequence
Secondary structure
-helix
-sheet
Tertiary structure
Quaternary structure
Subunits
Chaperones
Genetic code
Codons
Sense codons
Start codon
Nonsense codons
Stop or termination codons
Degenerate
Synonymous codons
Wobble base
Reading frame
Selenocysteine
Pyrrolysine
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Focal Points




The genetic code (Table 15.1)
The relationship between DNA, mRNA, tRNA, and polypeptide (Figure 15.3)
The directionality of polypeptide synthesis (Figure 15.4)
Levels of structures formed in proteins (Figure 15.7)
Exercises and Problems
For questions 1 to 6, match each of the following terms that are associated with the genetic code
to its correct definition. You may need to refer to Table 15.1 in the text for information.
_____ 1.
Universal
_____ 2.
Codons
_____ 3.
Start codon
_____ 4.
Stop codon
_____ 5.
Degenerate
_____ 6.
Wobble base
a.
b.
c.
d.
e.
f.
There are very few exceptions to the genetic code in living organisms.
A group of three nucleotides that specify a single amino acid.
The codon AUG.
The fact that the codons UAU and UAC code for the same amino acid.
The codon UAA.
There are more codons in the genetic code than there are amino acids.
For questions 7 to 12, select the level of protein structure that each statement refers to.
a. amino acid structure
b. primary structure
c. secondary structure
d. tertiary structure
e. quaternary structure
_____ 7.
α-helices and β-sheets
_____ 8.
The linear sequence of amino acids in a polypeptide
_____ 9.
R groups
_____ 10. Hydrophobic and ionic interactions
_____ 11. A carboxyl and amino functional group
_____ 12. The interaction of two or more polypeptides
For questions 13 to 15, complete the sentence with the most appropriate term(s):
13. The 21st and 22nd amino acids are ________ and ________.
14. ________ are proteins that bind to polypeptides and facilitate their proper folding.
15. Codons that specify the same amino acids are termed ________ codons.
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15.3 Experimental Determination of the Genetic Code
Overview
This section takes a close look at the experimental approaches that helped to deduce the
genetic code. The discussion begins with Francis Crick’s studies of frame-shift mutations in
bacteriophage T4 of E. coli. These experiments indicated that the genetic code is read in multiples
of three nucleotides (Refer to Table 15.3). The work of Marshall Nirenberg, Severo Ochoa, and
H. Gobind Khorana is covered next. They used synthetic RNA and a cell-free translation system
to decipher the genetic code (Refer to pages 342-345). The section ends with a discussion of other
experiments that helped decipher the genetic code, namely the synthesis of copolymers by
Khorana and colleagues (see Table 15.4), and the triple-binding assay of Nirenberg and Linder
(see Figure 15.9).
Outline of Key Terms
Frameshift mutation
Cell-free translation system
Focal Points



Evidence that the genetic code is read in triplets (Table 15.3)
Synthetic RNA helped to determine the genetic code (pages 342-345)
The triplet-binding assay (Figure 15.9)
Exercises and Problems
For questions 1 to 4, complete the following sentences with the most appropriate term(s):
1. Francis Crick performed experiments on ________ which indicated that the genetic code is
read in triplets.
2. The gain or loss of one or two bases alters the reading frame beyond the point of change,
thus causing a ________ mutation.
3. An in vitro translation system is also called a ________.
4. Synthetic mRNAs can be made using the enzyme ________.
For questions 5 to 8, refer to the following information: A researcher mixes together A, G, and C
ribonucleotides in a ratio of 3A:2G:1C. A synthetic RNA is then made from this mixture.
5. How many different codons can this mRNA contain?
6. Can this mRNA contain a start codon? A stop codon? Why or why not?
7. What proportion of the codons is expected to be 5′ AGC 3′?
8. What proportion of the codons is expected to be 5′ GCG 3′?
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15.4 Structure and Function of tRNA
Overview
The adaptor hypothesis was proposed in the 1950s by Francis Crick. It suggests that
tRNA molecules recognize codons in mRNA and carry amino acids to the ribosome. This section
of the chapter examines the structure and function of tRNA molecules.
The anticodon of a tRNA is complementary to a codon in mRNA (see Figure 15.10).
Transfer RNAs are the translators of the genetic code and so it is important to know their
structure (Figure 15.11). Aminoacyl-tRNA synthetases are a group of enzymes that attach the
correct amino acid to the tRNA. This process is called charging of the tRNA (Figure 15.12). The
final part of this section examines the wobble rules, which allow mismatches at the third position
in codon-anticodon pairing (Figure 15.13).
Outline of Key Terms
Anticodon
Wobble rules
Isoacceptor tRNAs
Adaptor hypothesis
Aminoacyl-tRNA synthetases
Aminoacyl tRNA
Charged tRNA
Focal Points




Recognition between tRNAs and mRNA (Figure 15.10)
Secondary structure of tRNA (Figure 15.11)
Catalytic function of aminoacyl-tRNA synthetase (Figure 15.12)
Wobble position and base-pairing rules (Figure 15.13)
Exercises and Problems
For questions 1 to 5, match the definition to its correct term.
______ 1. The enzyme that attaches the amino acids to the tRNA.
______ 2. According to the AT/GC rule, the complementary sequence to the codon.
______ 3. The idea that tRNAs recognize specific regions of the mRNA.
______ 4. The third base of the anticodon can tolerate mismatches.
______ 5. Two or more tRNAs that are recognized by the same codon.
a. aminoacyl-tRNA synthetase
b. adaptor hypothesis
c. wobble hypothesis
d. anticodon
e. isoacceptor tRNAs
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For questions 6 to 11, refer to the labels in the figure below.
______ 6. The label that indicates the location of the anticodon.
______ 7. The label that indicates the acceptor stem.
______ 8. The 5′ end of the tRNA.
______ 9. The site that will directly interact with the mRNA.
______ 10. The 3′ end of the tRNA.
______ 11. A charged tRNA will have an amino acid attached at this point.
15.5 Ribosome Structure and Assembly
Overview
Before proceeding to the stages of translation it is first necessary to study the ribosome,
which is the site of translation. A ribosome consists of a large and small subunit, each of which is
a composite of protein and ribosomal RNA (rRNA). The purpose of the ribosome is to provide a
workspace for the interaction of the mRNA and tRNA, and to ensure that the mRNA is being read
in the correct sequence.
Outline of Key Terms
Nucleolus
Ribosome
Ribosomal RNA (rRNA)
Aminoacyl (A) site
Peptidyl (P) site
Exit (E) site
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Focal Points


Composition of bacterial and eukaryotic ribosomes (Table 15.5)
Ribosomal structure (Figure 15.14)
Exercises and Problems
For questions 1 to 5, match the term with its correct definition.
______ 1. Ribosome
a. A site where the ribosomal subunits are assembled.
______ 2. P site
b. A site on the ribosome where uncharged tRNAs bind.
______ 3. E site
c. The site of translation.
______ 4. A site
d. The aminoacyl site.
______ 5. Nucleolus
e. The peptidyl site.
15.6 Stages of Translation
Overview
After introducing the genetic code, tRNAs and the ribosome, we are prepared to discuss
the process of translation. Note that translation involves three stages – initiation, elongation, and
termination. Although this is the same terminology as was used in transcription, the processes are
very different. Before entering into the details of the translation process, take a moment to review
Figure 15.15. Notice the interaction of the mRNA, ribosome, and tRNAs. Once you are familiar
with the overview, you can proceed into the more detailed examinations of protein synthesis.
During the initiation stage of bacterial translation, the mRNA and initiator tRNA
associate with the ribosomal subunits. The Shine-Dalgarno sequence promotes the binding of the
mRNA to the small ribosomal subunit (Figure 15.17). Elongation involves a repetitive cycle
during which successive amino acids are brought into the ribosome (via the A site) and linked
together with peptide bonds by peptidyl transferase (Figure 15.18). The growing polypeptide
chain is typically located in the P site, and uncharged tRNAs leave the ribosome through the E
site. During the termination stage, the stop codon in mRNA is reached and polypeptide synthesis
ends. Note that all stages of translation require protein factors, and energy in the form of GTP.
Outline of Key Terms
Coupling
Polyribosome
Polysome
Initiation
Initiator tRNA
Shine-Dalgarno sequence
Kozak’s rules
Elongation
Decoding function
Peptidyl transfer
Peptidyltransferase
Termination
Release factors
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Focal Points





Overview of translation (Figure 15.15)
The initiation (Figure 15.16), elongation (Figure 15.18), and termination (Figure 15.19)
stages of bacterial translation
Translational protein factors in bacteria and eukaryotes (Table 15.6)
Coupling between transcription and translation in bacteria (Figure 15.20)
Comparison of bacterial and eukaryotic translation (Table 15.7)
Exercises and Problems
For questions 1 to 7, indicate whether the statement is associated with prokaryotic (P) or
eukaryotic (E) translation, or both (B).
______ 1. Initiation factors are required.
______ 2. A 7-methylguanosine cap on the mRNA enhances initiation.
______ 3. The mRNA binds to the ribosomal subunit at the Shine-Dalgarno sequence.
______ 4. Sorting of the proteins occurs either during or after translation.
______ 5. Termination involves a single release factor.
______ 6. Transcription and translation occur simultaneously.
______ 7. Polyribosomes may occur on the mRNA.
For each of the following, indicate whether the statement is associated with initiation (I),
elongation (E), or termination (T) of translation.
______ 8. IF proteins stabilize the mRNA and ribosomal subunits.
______ 9. Nonsense codons enter into the A site.
______ 10. Release factors interact with stop codons.
______ 11. The ribosomal subunits associate with the mRNA.
______ 12. The ribosomal subunits dissociate.
______ 13. The peptidyl transfer reaction occurs between the amino acids of the P and A sites.
______ 14. The decoding function of translation occurs.
______ 15. Translocation of the ribosome occurs.
For each of the following, indicate whether the statement applies to the aminoacyl (A), exit (E),
or peptidyl (P) site of the ribosome.
______ 16. The initiator tRNA enters through this site.
______ 17. All other charged tRNAs enter through this site.
______ 18. Uncharged tRNAs leave through this site.
______ 19. Translation ends when a stop codon appears in this site.
______ 20. The site that is located in the middle of the ribosome.
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Chapter Quiz
1. Hydrophobic and ionic interactions between the R groups of amino acids occur at which level
of protein structure?
a. primary
b. secondary
c. tertiary
d. quaternary
2. The anticodon is located on the _________.
a. ribosome
b. mRNA
c. tRNA
d. rRNA
3. The molecule that attaches the amino acid to the tRNA is called _________.
a. peptidyltransferase
b. release factor
c. aminoacyl-tRNA transferase
d. aminoacyl-tRNA synthetase
4. In which of the following organisms can simultaneous transcription and translation occur?
a. prokaryotes
b. eukaryotes
c. both a and b
d. neither a nor b
5. Translocation of the ribosome occurs during _________.
a. elongation
b. termination
c. initiation
d. transcription
6. The Shine-Dalgarno sequence is involved in what stage of translation?
a. Initiation in bacteria
b. Elongation in bacteria
c. Initiation in eukaryotes
d. Termination in eukaryotes
e. none of the above
7. Which of the following is NOT a stop codon in most species?
a. UAA
b. UAC
c. UGA
d. UAG
8. Which of the following statements regarding the genetic code is FALSE?
a. It is nearly universal.
b. It is degenerate.
c. It allows wobble in the third base.
d. All of the above are correct.
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9. If the genetic code was a quadruplet (not triplet) code, how many codons would you expect?
a. 64
b. 128
c. 256
d. 512
10. During the peptidyl transferase reaction, the ribosomal ______ receives the polypeptide chain.
a. A site
b. P site
c. E site
d. S site
Answer Key for Study Guide Questions
This answer key provides the answers to the exercises and chapter quiz for this chapter. Answers
in parentheses ( ) represent possible alternate answers to a problem, while answers marked with
an asterisk (*) indicate that the response to the question may vary.
15.1
1. Garrod
2. alkaptonuria
3. homogentisic
4. Beadle and Tatum
5. Neurospora crassa
6. polypeptide
7. protein
15.2
1. a
2. b
3. c
4. e
5. f
6. d
7. c
8. b
9. a
10. d
11. a
12. e
13. selenocysteine and pyrrolysine
14. chaperones
15. synonymous
15.3
1. bacteriophages T4
2. frameshift
3. cell-free translation system
4. polynucleotide phosphorylase
5. 33 = 27
6. No and No; because there are no U’s in the mixture
7. (3/6)(2/6)(1/6) = 6/216 or 2.78%
8. (2/6)(1/6)(2/6) = 4/216 or 1.85%
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15.4
1. a
2. d
3. b
4. c
5. e
6. d
7. c
8. b
9. d
10. a
11. a
15.5
1. c
2. e
3. b
4. d
5. a
15.6
1. B
2. E
3. P
4. E
5. E
6. P
7. B
8. I
9. T
10. T
11. I
12. T
13. E
14. E
15. E
16. P
17. A
18. E
19. A
20. P
1. c
2. c
3. d
4. a
5. a
6. a
7. b
8. d
9. c
10. a
Quiz
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