Protein translation and post-translational modification

Question 1

How does a ribosome translate mRNA into a protein sequence?

Answer 1

Ribosome scans from 5’ end of mRNA starting at first AUG until it reaches the first stop codon in frame
Initiation
Elongation
Termination

Question 2

What is the start codon and its corresponding amino acid?

Answer 2

AUG - Methionine

Question 3

What are the stop codons?

Answer 3

UAA, UAG, UGA

Question 4

What is the structure of mRNA?

Answer 4

At the 5’ end, mRNA has a 7-methyl guanosine cap followed by a 5’ untranslated region before the start codon
mRNA ends with the 3’ untanslated region and a poly-A tail (prevents degradation)

Question 5

How is an amino acid bound to a tRNA molecule?

Answer 5

Aminoacyl tRNA synthetase uses ATP to bind an amino acid to itself to form an E-AMP-Amino Acid (adenylated amino acid)
When the amino acid binds to the 3’ end of tRNA, AMP and the enzyme dissociates

Question 6

What comprises the pre-intiation complex?

Answer 6

40S ribosomal unit
Met-tRNA
eIF2 (initiation factor)
GTP

Question 7

How is the pre-initiation complex assembled during initiation?

Answer 7

Subunits dissociate
Assembly of the pre-initation complex:
-Eukaryotic intiation factors 4E and 4G bind to 5’ cap
-These are recognised by 40S subunit, Met-tRNA and eIF2
mRNA binds to pre-initiation complex
mRNA is scanned for AUG start codon
GTP on eIF2 is hydrolysed, allowing the binding of the 60S subunit

Question 8

Describe the process of elongation

Answer 8

tRNA binds to amino acyl site A
Peptidyl transferase catalyses the formation of a peptide bond between two amino acids
tRNA is translocated to the P site while the first tRNA leaves

Question 9

What are elongation factors?

Answer 9

Proteins that promote the movement of the ribosome along the mRNA, using GTP
GTP hydrolysis enhances efficiency and accuracy of translation:
-provides pauses to allow incorrect base pairs to dissociate

Question 10

Describe the process of termination

Answer 10

Release factors bind to empty A site when a stop codon is reached
Allows peptidyl transferase to catalyse the transfer of the completed protein chain to water, releasing it from the ribosome
tRNA released, subunits dissociate and mRNA released

Question 11

Define polyribosome

Answer 11

Many ribosomes work on each strand at once, working at a speed of 15 amino acids per second

Question 12

Why do some antibiotics inhibit protein synthesis in prokaryotes but not in eukaryotes?

Answer 12

Ribosomes in prokaryotes are 70S, smaller than in eukaryotes, allowing for selective prokaryotic targeting
e.g Stroptomycin, Tetracycline, Erythromycinm Chloramphenicol, Puromycin

Question 13

What features must a newly synthesised protein have in order to enter the secretory pathway?

Answer 13

Signal sequence - the first 20-24 amino acids that state the destination of the protein, consisting of mainly hydrophobic amino acids

Question 14

How does a protein enter the ER?

Answer 14

Signal recognition particle recognises and binds to signal sequence, halting translation
SRP binds to SRP receptor on ER and translation resumes
This binding triggers delivery of polypeptide via a pore into the lumen of ER
Signal sequence then cleaved by signal peptidase

Question 15

What are the ways in which a newly synthesised protein can be post-translationally modified?

Answer 15

Formation of disulfide bonds

Addition of carb/lipid/phosphate etc.

Question 16

How is insulin modified?

Answer 16

Insulin translated to preproinsulin which has a signal sequence
Converted to proinsulin in the ER/Golgi with signal sequence removed
Disulfide bonds are formed before C Chain removed