8/15/14 - Protein Synthesis

Question 1

What is the purpose of the 7-methylguanosine cap in translation initiation?

Answer 1

It binds the eIF cap-binding complex, which facilitates the recruitment of the small ribosomal subunit to the mRNA transcript.

Question 1

What are the sizes of eukaryotic ribosomal subunits?

Answer 1

40S + 60S > 80S

Question 2

What are the 3 stop codons?

Answer 2

UAA, UAG, UGA

Question 4

What are 3 key features of the genetic code?

Answer 4

1. It is redundant (there is more than 1 codon for each amino acid) 2. It is universal 3. Some codons have additional functions (start/stop codons)

Question 5

List the 5 main steps of eukaryotic translation initiation

Answer 5

1. Initiation tRNA bound to small ribosomal subunit 2. eIF4 complex binds to 5’ cap, 3’ tail 3. Ribosomal subunit recognizes 5’ cap and associated eiF4 complex, binds 4. tRNA scans for the first AUG using ATP as energy 5. GTP hydrolysis of eIF2 facilitates large ribosomal subunit binding, translation can now begin

Question 6

What is the significance of the ester bond formed between an amino acid an tRNA?

Answer 6

It provides the energy for peptide bond formation during elongation.

Question 6

What happens to EFTu-GTP if the codon/anticodon base pairing is incorrect?

Answer 6

If the base pairing is incorrect, GTP hydrolysis will be delayed, giving time for the tRNA to dissociate from the ribosome.

Question 7

What are the main functions of the large and small ribosomal subunits?

Answer 7

Large: Catalyzes peptide bond formation Small: Provides framework to accurately match codon/anticodon

Question 7

Why are eukaryotes monocistronic?

Answer 7

mRNAs usually require a 5’ cap to bind the ribosomal subunit. Since there is only 1 5’ cap per mRNA, there can only be one region where translation is initiated.

Question 9

What is the difference between aminoacyl-tRNA synthetases in eukaryotes and prokaryotes?

Answer 9

Eukaryotes possess aminoacyl-tRNA synthetases specific to each amino acid. Prokaryotes possess aminoacyl-tRNA synthetases that recognize more than 1 amino acid.

Question 10

What are the sizes of prokaryotic ribosomal subunits?

Answer 10

30S + 50S > 70S

Question 11

What effect might a deletion/insertion have on protein synthesis?

Answer 11

Frameshift mutation

Question 12

What are two ways aminoacyl-tRNA synthetase recognizes the correct tRNA?

Answer 12

1. Anticodon recognition (does not work when multiple codons code the same amino acid) 2. Structural recognition at certain regions in the tRNA

Question 13

Describe what occurs in A, P, and E sites during elongation?

Answer 13

A: tRNA enters and base pairs with mRNA P: Peptide bond is formed between amino acids at the A and P site. (also the initiation site) E: tRNA exit site

Question 14

What are two ways aminoacyl-tRNA synthetase recognize the correct amino acid?

Answer 14

1. Unique conformation of the binding site promotes correct binding of the amino acid (ex. a large amino acid won’t fit in a small binding site). 2. aminoacyl-tRNA synthetase pushes bound amino acid into editing site; if it fits, it is the incorrect amino acid

Question 15

What is the main function of EFTu-GTP?

Answer 15

It binds loaded tRNA as it enters the ribosome to ensure the correct tRNA has been recruited.

Question 16

What are 4 differences between transcription and translation in prokaryotes?

Answer 16

1. Transcription/translation can occur simultaneously 2. mRNAs are not processed with 5’ caps and polyA tails. 3. There is only 1 RNA polymerase 4. Ribosomal subunits are different sizes

Question 17

What is the main function of the Shine-Dalgarno box in prokaryotes?

Answer 17

SD box/16S rRNA base pairing determines the correct reading frame for translation.

Question 17

What are the two mechanisms by which tRNA is moved from A > P > E sites during elongation?

Answer 17

1. Large ribosomal subunit translocates relative to the small subunit, placing the tRNAs in hybrid A/P, P/E sites. 2. EFG-GTP is hydrolized to induce a conformational change that pushes the tRNAs fully into the P and E sites.

Question 19

What are the major steps in protein synthesis?

Answer 19

1. Loading (or charging) the tRNA with an amino acid 2. Initiation 3. Elongation 4. Termination

Question 20

What is the sequence for a start codon?

Answer 20

AUG

Question 21

What amino acid does a start codon encode?

Answer 21

Methionine

Question 22

List the 3 main steps of prokaryotic translation initiation

Answer 22

1. Small ribosomal subunit binds at the SD box on the mRNA 2. tRNA/F-Met/IF2-GTP complex binds to the initiation site. 3. IF2-GTP is hydrolyzed to facilitate assembly of the large ribosomal subunit, initiation factors dissociate and translation can now begin

Question 22

How is eIF2-GDP catalyzed back into GTP form?

Answer 22

eIF2B (a guanine nucleotide exchange factor)

Question 24

Why are eukaryotes polycistronic?

Answer 24

The 5' cap is insignificant in prokaryotes, since the ribosomes bind the mRNA through the activity of Shine-Dalgarno sequences. There can be multiple SD boxes on each mRNA, so more than 1 protein can be encoded.

Question 25

How would the inactivation of eIF2B alter protein synthesis?

Answer 25

eIF2B activates eIF2, which binds the large ribosomal subunit to the small subunit in order to begin translation. Inactivating eIF2B would stop protein synthesis at the initiation stage.

Question 27

What happens to EFTu-GTP is the codon/anticodon base pairing is correct?

Answer 27

GTP is hydrolized and EFTu-GDP dissociates from the tRNA, which allows the tRNA to fully move into the A site where translation can proceed. A 2nd delay following GTP hydrolysis offers 1 more opportunity for an incorrect tRNA to dissociate from the ribosome.

Question 28

What effect will a frameshift mutation have on protein synthesis?

Answer 28

The mRNA will be translated along the wrong reading frame.

Question 29

Describe the mechanism of action for Puromycin

Answer 29

It has similar structure to tyrosyl-tRNA, and can be incorporated into the growing polypeptide chain during elongation. However, further elongation is terminated due to its lack of a high energy bond.

Question 30

What effect will a nonsense mutation have on protein synthesis?

Answer 30

An early stop codon will result in a truncated protein.

Question 31

What is the main function of EFG?

Answer 31

It binds near the A site on the ribosome and accelerates the movement of tRNAs to the P and E sites.

Question 32

Describe the wobble phenomenon and how it relates to the redundancy of the genetic code

Answer 32

The 5' base of the anticodon is not confined to a single nucleic acid for each amino acid. In other words, more than 1 nucleic acid at this position can code for the same amino acid.

Question 33

Which steps in protein synthesis is GTP hydrolysis thought to occur?

Answer 33

Initiation (EFTu/EFG) Elongation (eIF2)

Question 34

Draw the interaction between a messenger RNA codon and its cognate tRNA, labeling the orientations of both RNA strands.

Answer 34