Exam 2- Translation

Question 1

What are codons?

Answer 1

A set of three adjacent nucleotides called a triplet
Nucleotide options- A, U, G, or C
Total of 61 possible codons to code for 21 amino acids
Are very specific (unambiguous)

Question 2

Describe start and stop codons.

Answer 2

Start Codon: AUG

• only codon that codes for methionine
• towards the 5’ end of DNA

Stop Codons: UGA, UAG, UAA
- Only exception in mitochondrial DNA, where UGA codes for Tryptophan

Question 3

What are the 5’ and 3’ untranslated regions (UTR’s)?

Answer 3

5’UTR: Regulates translational efficiency

3’UTR: Controls mRNA stability. Stabilizing and destabilizing proteins bind to this end

Question 4

A codon is “universal” and often degenerate. Explain.

Answer 4

Universal: they code for the same amino acid across multiple organisms with few exceptions.
Ex. AUG always codes for methionine

Degenerate: multiple codons can code for the same amino acid
Ex. GUU, GUC, GUA, and GUG all code for Valine

Codons are always specific to an amino acid

Question 5

Describe Huntington Disease.

Answer 5

Insertion of a CAG codon in tandem repeats in the Huntington gene

Results in Huntington protein with several glutamine residues that accumulates and causes defects

Advanced stage symptoms include uncoordinated, jerky body movements, unsteady gait, mood change, and higher order cognition defect

Question 6

Missense Mutation

Answer 6

Point mutation in which a single nt change results in a codon that codes for a different amino acid

Ex. Sickle cell anemia: single nt causes Glu to Val resulting in defective beta globulins

Question 7

Nonsense mutation

Answer 7

A point mutation that results in a premature stop codon

Question 8

Silent mutation

Answer 8

Mutation in a codon that results in the same amino acid and no observable effect
Ex. UCA (serine) —> UCU (serine)

Question 9

Addition mutation

Answer 9

Addition of one or more nucleotides to a DNA sequence

Question 10

Deletion mutation

Answer 10

Deletion of one or more nucleotides in a DNA sequence

Question 11

Frame shift mutation

Answer 11

Insertion or deletion of 1 or 2 nt’s leading to a reading frame change and may lead to a truncated protein if causes a stop codon somewhere down the line

Note- loss or insertion of 3 does not alter reading frame
Ex. Cystic fibrosis loss of 3 nt’s causes loss of Phe in CFTR protein

Question 12

Describe the tRNA synthetase catalyzed reaction of aminoacyl-tRNA formation

Answer 12

TRNA synthetase- is an enzyme that attaches the appropriate amino acid onto its tRNA

In initiation, tRNA synthetase charges an amino acid onto tRNA

• amino acid + ATP + amino acyl tRNA synthetase (E) —> E-AMP-amino acid (enzyme-substrate complex)
• E-AMP-amino acid + tRNA —> amninoacyl tRNA bound to amino acid + AMP + E
• carboxyl group of amino acid is now bound to the 3’ end of tRNA!

Question 13

Describe the subunits of prokaryotic and eukaryotic ribosome complex.

Answer 13

Prokaryotic Ribosome:
50S (contains 5S and 28S RNA and 31 proteins)
30S (contains 16S RNA and 21 proteins)
Total Ribosome = 70S

Eukaryotic Ribosome:
60S large subunit (contains 5S, 5.8S, 28S, and 50 proteins)
- catalyzes formation of peptide bonds that link amino acids
40S small subunit (contains 18S RNA and 30 proteins)
Total Ribosome = 80S

Question 14

The three sites of ribosome that is involved in translation/protein synthesis

Answer 14

E= Empty tRNA site for exit 
P= peptidyl tRNA (already extended, attached to a growing chain)
A= incoming aminoacyl tRNA binding site

Question 15

What are polysomes?

Answer 15

Cluster of ribosomes held together by a strand of messenger RNA that each ribosome is translating

Question 16

Explain the functions of RER mediated protein synthesis.

Answer 16

RER has ribosomes studded on it
RER ribosomes make proteins for ER lumen, Golgi, Lysosomes, and Plasma membrane

Cytosolic ribosomes make proteins for cytosol, nucleus, mitochondria, and peroxisome

Question 17

Three steps involved in protein synthesis

Answer 17

Initiation, Elongation, Termination

Question 18

Explain Initiation

Answer 18

• mRNA associates with small ribosomal subunit and requires initiation factors
• charging of amino acid onto tRNA

In prokaryotes, Shine Delgado sequence (purine rich) binds 16S RNA and the whole mRNA aligns in position for translation. Fmet-RNA is the initiating tRNA

In eukaryotes, 5’-Cap is uses as an aligning factor with small subunit. Met-tRNA is the initiating tRNA

Question 19

Elongation

Answer 19

Requires elongation factors that assist the peptide bond formation between carboxyl group of first aa and amino group of 2nd aa

The 28S RNA is catalytic (ribozyme), peptidyl transferase that catalyze the formation of peptide bonds

Question 20

Termination

Answer 20

Involves release factors and a stop codon

Question 21

What are the steps that use GTP and ATP during protein synthesis ?

Answer 21

Cleavage of 4 high energy bonds per 1 amino acid attached to the growing polypeptide chain

2 ATP for aminoacyl tRNA synthesis
1 GTP for aminoacyl tRNA to bind to A site
1 GTP for translocation

And at the end,
1 GTP for termination

Question 22

Puromycin

Answer 22

Aminoacyl tRNA analog

Inhibits growth of both prokaryotic and eukaryotic proteins

Question 23

Chloramphenicol

Answer 23

Prokaryotic peptidyl transferase (E that forms peptide bonds between adjacent aa’s using tRNA’s during translation) inhibitor

Question 24

Tetracycline

Answer 24

Interacts with 30S subunit and blocks access of aminoacyl tRNA to the mRNA ribosome complex

Question 25

Streptomycin

Answer 25

Interferes with initiation by binding to 30S

Question 26

Clindamycin

Answer 26

Binds to 50S ribosome and inhibits elongation

Question 27

Erythromycin

Answer 27

Binds to 50S ribosome and inhibits elongation

Question 28

Diphtheria GToxxin

Answer 28

Inactivates eukaryotic elongation factor eEF-2 and prevents translocation

Question 29

_____________ has its own ribosome with a lot of similarities to prokaryotic ribosomes, thus side effects can be seen in some prokaryote targeted drugs due to the cross reactivity with mitochondrial ribosomes

Answer 29

Mitochondria