HRR: translation

Question 1

Compare and contrast the main structural components of prokaryotic versus eukaryotic mRNAs.

Answer 1

Prokaryotes: 70s with 50s and 30s subunits; 50s has 23 and 5s rRNAs, 30 has 16s rRNAs
Eukaryotes: 80s with 60 and 40s subunits; 60 has 28, 5, and 5.8s rRNAs, 40 has 18s rRNAs

Question 2

Describe secondary and tertiary structure of the tRNA molecule and pinpoint the anticodon region and the amino acid attachment site

Answer 2

Secondary: a clover leaf structure; base of the clover holds the anticodon. 3’ end has CAA, and the last A attaches the amino acid.

Tertiary: an L shaped structure; the anticodon is at the base of the structure. 3’ end has CAA, and the last A attaches the amino acid.

Question 3

Describe a “charged” tRNA molecule and how it is generated.

Answer 3

A tRNA molecule with the amino acid attached via an aminoacyl linkage. Once charged, it can go to the ribosome and deliver the amino acid.

Question 4

Describe the function of aminoacyl-tRNA synthetases in implementing the genetic code.

Answer 4

Aminoacyl-tRNA synthetases recognize their corresponding amino acid and catalyze the reaction in which it is attached correctly to the corresponding tRNA. There is an editing function if an incorrect aminoacyl linage is formed.

Question 5

Describe how aminoacyl synthetases work

Answer 5

They recognize the R group on an amino acid and utilize ATP to add AMP on the carboxyl group, forming an aminoacyl intermediate. It then finds the correct tRNA and switches the AMP for the tRNA (CAA end).

Question 6

Explain why the genetic code is “degenerate”.

Answer 6

More than one codon can code for the same amino acid

Question 7

What are the stop codons? Start codon?

Answer 7

UAG, UAA, UGA; AUG

Question 8

Describe codon-anticodon base pairing and indicate the “wobble” bases.

Answer 8

Codons (read 5’ to 3’) are complementary to anticodons (read 3’ to 5’)

Wobble bases are the third bases of codons and the first bases of anticodons, and they are what allows for degeneracy.

Question 9

Explain why inosine is a frequent base modification in the anticodon loop of tRNA.

Answer 9

Deamination of adenosine forms inosine; inosine base pairing maximizes the number of codons that a tRNA can read by forming non-standard base pairings.

Question 10

Define the Shine-Dalgarno sequence and describe its role in translational initiation

Answer 10

This is how the ribosome recognizes where the start codon is in prokaryotes. The mRNA has this sequence upstream of the start codon and it functions as a marker for the recognition of the codon. The 30s subunit binds to the sequence via complementary base pairing.

Question 11

Compare the initiation step of translation in prokaryotes versus eukaryotes. Describe how the mechanisms for recognition of the AUG start codon differ

Answer 11

Prokaryote: uses the shine-delgarno sequence
Eukaryote: uses linear scanning to find the Kozak sequence that contains the start codon

Question 12

Describe initiation of translation in prokaryotes

Answer 12

-IF1 and IF3 bind to 30s unit
-IF2-GTP binds with the charged initiator tRNA, fMet-tRNA
-The IF1 and IF3 dissociate
-The 50s unit joins
-GTP hydrolyzes, resulting in the release of IF2-GDP

Question 13

Describe initiation of translation in eukaryotes

Answer 13

-Formation of the ternary complex containing met-tRNA-EIF2-GTP
-The ternary complex and eukaryotic initiation factors (namely EIF3 and EIF4) bind to the 40s subunit. This forms the preinitiation complex.
-EIF-4F helps the PIC bind to the m7Gppp cap on the 5’ end of mRNA. The three main subunits are 4G, 4E 44G connects to EIF3. Associated with 4G are 4A (RNA helicase) and 4E (binds the cap)
-PIC finds start codon
-EIF5 triggers hydrolysis of EIF2-GDP, thereby releasing them from the ribosome. This enables the 60s unit to bind to form the 80s initiation complex.

Question 14

Describe the modification of methionine on the initiator in bacteria

Answer 14

They have an enzyme that protects the amino group, because if not it can interact with other parts of the peptide. The enzyme formylates it.

Question 15

Specify the roles of the following elongation factors: EF-Tu, EF-Ts, EF-G during the
elongation phase of translation. Specify their counterparts in eukaryotes.

Answer 15

EF-Tu: binds to charged tRNA and brings it to the A site to base pair and form peptidyl tRNA; functions in proofreading. eEF-1a in eukaryote
EF-Ts: exchanges GDP-pi for GTP to activate EF-Tu/eEF-1a; eEF-1bg in eukaryote
EF-G: catalyzes the translocation of peptidyl tRNA from the A site to the P site; eEF-2 in eukaryote

Question 16

Specify two steps in the elongation phase of translation that require GTP hydrolysis.

Answer 16

Attaching the codon to anticodon, moving peptidyl tRNA to the P site

Question 17

Name the enzyme that catalyzes peptide bond formation.

Answer 17

Peptidyl transferase

Question 18

Describe how proofreading occurs during the elongation step of translation.

Answer 18

GTPase on EF-Tu will not fire off if the base-pairing is not correct. It gets kicked out before the GTPase fires off if it is not correct.

Question 19

mRNA is translated in the ___ direction and that proteins are synthesized
from the ___ end to the ____

Answer 19

5 to 3; amino to carboxyl

Question 20

Define polyribosomes and explain how they improve the efficiency of translation.

Answer 20

Multiple ribosomes translating a single mRNA molecule; more ribosomes per mRNA molecule leads to a higher efficiency

Question 21

Describe the functions of protein chaperones

Answer 21

Facilitate protein folding by binding and stabilizing polypeptides; allows for proper conformational forming; there are co-translational chaperones such as heat shock proteins

Question 22

Describe the mechanism by which translation is terminated.

Answer 22

-Stop codon moves to the A site
-Release factors are attracted and recognize the stop codon
-The RF binds to the A site and hydrolyze the tRNA from the C terminal AA of the polypeptide chain

Question 23

Describe how miRNAs are generated and the mechanisms by which miRNAs regulate
gene expression

Answer 23

miRNAs genes are transcribed by Pol II and made inter pri-miRNA transcripts. They are processed and into small double stranded RNAs that can be recruited into RISC complexes. The RISC separates the strands and helps a strand base pair with the target strand. This allows for regulation via stopping translation or triggering degradation of the protein.

Question 24

The UTR is part of the ___ (intron or exon)

Answer 24

exon

Question 25

In prokaryotes, transcription occurs ___ translation

Answer 25

Simultaneously with; mRNA is not processed or spliced

Question 26

What is a p-body?

Answer 26

Storage depots for mRNA; non-membranous cytosolic organelles