Ch. 18 Protein Synthesis

Question 1

What are the prokaryotic and eukaryotic ribosomes and how are they similar?

Answer 1

70S = prokaryotic and 80S = eukaryotic. They are structurally similar.

Question 2

What is the small ribosomal subunit? Prokaryotic vs Eukaryotic?

Answer 2

Where the tRNA interacts with mRNA. 30S = prokaryotic and 40S = eukaryotic.

Question 3

What is the large ribosomal subunit? Prokaryotic vs eukaryotic?

Answer 3

Where peptidyl transferase makes peptide bonds. 50S = prokaryotic and 60S = eukaryotic.

Question 4

What is each ribosomal subunit made up of? (2)

Answer 4

rRNA and protiens

Question 5

How is ribosomal RNA described across all domains of life? What does this indicate (2)?

Answer 5

Ribosomal RNA is highly conserved among domains of life. This indicates a fundamental role in ribosome function and an ancient origin in cells.

Question 6

What is responsible for the ribosome function?

Answer 6

RNA is primarily responsible for ribosome functions, not proteins.

Question 7

How do eukaryotic ribosomes differ from prokaryotes? (3)

Answer 7

They are larger, more complex, and cannot spontaneously form in vitro. This suggests a complex assemble process.

Question 8

What are the “big picture” steps of polypeptide synthesis? (3)

Answer 8

Initiation, Elongation, Termination.

Question 9

What are the three binding sites in the small ribosomal subunit?

Answer 9

A site: aminoacyl-tRNA binding site
P site: peptidyl-tRNA binding site
E site: exit/eject site

Question 10

What does the large subunit catalyze? (2)

Answer 10

1. Breaking of the bond between the amino acid and the tRNA in the P site
2. Formation of a peptide bond between that same amino acid and the amino acid in the A site

Question 11

What are the steps of tRNA charging? (3)

Answer 11

1. amino acid carboxyl group attacks the ɑ phosphate of ATP to produce aminoacyl-AMP
2. aminoacyl-AMP reacts with the 3’ end of the tRNA
3. a high energy bond is produced that provides energy for peptide bond formation

Question 12

What are class I aminoacyl-tRNA synthetases?

Answer 12

They add the amino acid to the 2’OH of ribose, which then gets moved to the 3’OH.

Question 13

What are class II aminoacyl-tRNA synthetases?

Answer 13

They add the amino acid to the 3’OH of ribose.

Question 14

Where are the differences in Prokaryotic and Eukaryotic translation? (3)

Answer 14

Differences in initiation, elongation, and termination. Activation of amino acids and posttranslational processing are the same.

Question 15

What is the generic process of translation initiation? (2)

Answer 15

1. small subunit and initiator tRNA associate with the mRNA
2. large subunit binds to complete the ribosome assembly

Question 16

How is prokaryotic translation initiation started?

Answer 16

A 16S rRNA recognized the Shine-Delgarno sequence (ribosome binding site) about 10 bp upstream of an AUG.

Question 17

How are ribosome binding sites structured in a chromosome?

Answer 17

Genes may each have their own RBS, or, the stop codon of one reading frame can overlap with the start codon of another reading frame.

Question 18

How is the next sequence/gene translated?

Answer 18

The ribosome will shift the reading frame to translate the next sequence.

Question 19

What are the steps of prokaryotic translation initiation after it is started? (5)

Answer 19

The 70S assemble requires initiation factors (IFs)
1. IF1 binds the A site and prevents tRNAs from binding the A site
2. IF3 binds the E site and prevents the large subunit from binding
3. mRNA binds the 30S/IF1/IF3 complex with the start codon in the P site
4. IF2 binds and loads the initiator tRNA into the P site
5. the large subunit binds to complete the 70S ribosome

Question 20

What are the steps of eukaryotic translation initiation? (6)

Answer 20

1. IFs associate with the 40S small subunit
2. additional IFs load the initiator tRNA into the P site of the 40S
3. mRNA is loaded based on recognition of the 5’ cap by elF4F
4. mRNA is fed through the complex and scanned for a start codon
5. the large subunit binds to form the 80 S ribosome
6. multiple elF4’s cause circularization of the mRNA

Question 21

Why does circularization occur in eukaryotic translation initiation?

Answer 21

Circularization of the mRNA is due to interactions with poly(A) binding protein at the poly(A) tail. It facilitates translational regulation.

Question 22

What is an advantage of circularization in eukaryotes?

Answer 22

Multiple ribosomes can bind at the 5’ cap and assemble to translate the circularized mRNA.

Question 23

Whaat is the basic idea of translation elongation in both prokaryotes and eukaryotes? (2)

Answer 23

Amino acids are added one at a time. It requires three elongation factors (EFs) that have different names in prokaryotes and eukaryotes, but the same function.

Question 24

What is the general sequence of events during translation elongation? (5)

Answer 24

1. charged tRNA enters A site
2. a.a. in P site is added to the a.a. in the A site
3. ribosome shifts one codon
4. uncharged tRNA exits from E site
5. new, charged tRNA enters empty A site

Question 25

What is EF-Tu? EF-1ɑ?

Answer 25

EF-Tu binds GTP and aminoacyl-tRNA, and loads the tRNA into the ribosome. EF-1ɑ is the eukaryotic equivalent.

Question 26

How does the correct aminoacyl-tRNA enter the ribosome?

Answer 26

codon/anticodon interaction ensures the aminoacyl-tRNA is correct.

Question 27

What happens to EF-Tu﹣GTP once bound to ribosome? (2)

Answer 27

GTP is hydrolyzed and EF-Tu﹣GDP is released. EF-Tu﹣GDP will be regenerated to EF-Tu﹣GTP by EF-Ts (EF-1β is the eukaryotic equivalent).

Question 28

What are the steps of the peptidyl transferase reaction? (3)

Answer 28

1. the amine group of the a.a in the A site is a nucleophile that attacks the carbonyl carbon of the a.a. in the P site 2. bond rearrangements lead to a peptide bond and both a.a's are in site A 3. the tRNA in the P site is uncharged (deacylated)

Question 29

What happens when a peptide bond forms? (4)

Answer 29

1. once a peptide bond forms, the ribosome translocates one codon towards the 3' end of the mRNA 2. the dipeptidyl tRNA shifts from the A site to the P site 3. the deacylated tRNA shifts from the P site to the E site 4. the third codon is now in the A site

Question 30

What is EF-G and what does it do? EF-2? (3)

Answer 30

Translocation depends on GTP hydrolysis by EF-G (EF-2 is the eukaryotic equivalent). 1. EF-G﹣GTP binds the sarge subunit A site and causes it to shift one codon to the 3' end (misaligned) 2. GTP hydrolysis to GDP leads to the small subunit shifting one codon (realigned)

Question 31

Once the third amino acid of a polypeptide is added, what is the general translation elongation process? (3)

Answer 31

1. with the A site now empty, a new tRNA-aa can bind 2. the process of new peptide bond formation and ribosome shifting repeats 3. peptide bond formation will continue with charged tRNAs entering A site, but will terminate when a stop codon enters the A site

Question 32

How does translation terminate for all organisms?

Answer 32

Release factors (RFs) terminate translation.

Question 33

What is the process of prokaryotic translation termination? (4)

Answer 33

1. RF-1 (UAG, UAA) or RF-2 (UGA, UAA) bind a stop codon in the A site 2. binding of the RF causes peptidyl transfer of the polypeptide to water instead of an a.a. 3. RF-3 hydrolyzes GTP and catalyzes the removal of RF-1/2 4. ribosomal subunits are then recycled

Question 34

How does eukaryotic translation termination differ from prokaryotic?

Answer 34

They have a similar mechanism, but they only have one RF that recognizes all stop codons.