Protein Translation and Post-Translational modification Flashcards
5’ 7-methyl guanosine cap of mRNA
entry site for ribosome
5’ UTR of mRNA
untranslated region (non-coding region) may be involved in regulation of translation or mRNA stability
What follows the 5’UTR on mRNA?
Coding region
3’ UTR
PolyA tail
What is the function of the polyA tail on mRNA?
To protect the 3’end from degradation
Why is the genetic code described as degenerate?
Common AAs have several codons
What are the stop codons?
UAA, UAG, UGA
What is the Methionine codon?
AUG
What is the significance of Methionine?
It is the 1st AA in nearly all polypeptides
Transfer RNA function
Transport AA’s to ribosome
How many tRNA per AA?
> ,1
Where is an AA attached to on tRNA
3’ end
What enzyme is involved in the transfer of an AA to tRNA?
Aminoacyl tRNA synthetase
How many Aminoacyl tRNA synthetases per AA and why is this important?
1 per AA
Important in fidelity of translation - ensure correct AA is bound to correct tRNA
How does Aminoacyl tRNA synthetase work?
Aminoacyl tRNA synthetase gets activated (by ATP hydrolysis) to form adenylated AA which binds with a molecule of AMP and binds to the relevant AA
This adenylated AA complex then binds to the tRNA and transfers the AA to the 3’ end of the tRNA
The adenylated AA and AMP dissociate
What are the 3 stages of translation?
Initiation
Elongation
Termination
Initiation in translation
Dissociation of ribosome subunits (eukaryotic 40S + 60S)
Assembly of preinitiation complex containing Met-tRNA + eukaryotic translation initiation factors (eIFs) + 40S subunit
Binding of mRNA to preinitiation complex
Binding of 60S subunit
Elongation in translation
Binding of new tRNA to immediately adjacent A site on ribosome in frame with initiator Met
Catalysis of peptide bond formation between the 2 amino acids by peptidyl transferase (PT) on 60S subunit
Translocation of tRNA to P site and dissociation of 1st tRNA
Ribosome slides along so previous tRNA is now in the P site and the used tRNA falls off- facilitated by Elongation Factors (EFs) using GTP
Why do elongation factors use GTP?
To enhance efficiency and accuracy of translation by providing “pauses” (e.g. GTP hydrolysis) that allow incorrect base pairs to dissociate
Termination in translation
Recognition of stop codon
Release of peptide chain
Dissociation of release factors and ribosomes
What occurs in recognition of stop codon in termination of translation?
Release factors (proteins) bind to empty A site (no tRNA for stop codons) Peptidyl transferase catalyses transfer of the completed protein chain to water (hydrolysis) and releases it from the ribosome
Polyribosome
several ribosomes work on each strand at once
Allows much more protein to be produced in the same amount of time
Many antibiotics inhibit protein synthesis in prokaryotes
Antibiotics exploit differences between prokaryotic and eukaryotic ribosomes and translation factors
Allows selective inhibition of prokaryotes
What does Streptomycin inhibit?
Inhibits initiation
What does Tetracycline inhibit?
Inhibits AA-tRNA binding
What does Erythromycin inhibit?
Inhibits translocation
What does Chloramphenicol inhibit?
Inhibits peptidyl transferase
What does Puromycin do?
Terminates elongation prematurely
Where does protein synthesis occur and why do cells need a mechanism to transfer proteins across membranes?
Occurs in Cytoplasm
Cellular organelles are bound by a membrane
Where are secretory and transmembrane proteins synthesised?
In rough endoplasmic reticulum
Signal sequence
Proteins destined to be secretory or transmembrane have a special sequence (1st 20-24 AAs)
Found at N terminus of polypeptide
Rich in hydrophobic amino acids (lipophilic so can cross membranes)
Describe how proteins enter the RER
The signal sequence binds to a signal recognition particle (SRP) and this binding halts translation
The SRP then binds to an SRP receptor on the RER membrane and translation resumes
The binding of SRP to the SRP receptor triggers the assembly of a protein channel through which the polypeptide is threaded into the lumen of the RER
What extra feature do proteins that are destined to be transmembrane have?
They encode 1 or more extra hydrophobic sequences that hold them in the membrane
What happens once the protein has entered the RER?
The signal sequence is cleaved by signal peptidase and is degraded.
Result: main protein, which is folded.
Why are most proteins further modified before becoming fully functional?
Only 20 amino acids
Cell uses post-translational modifications to increase diversity
Give 3 examples of post-translational modifications
Disulphide bond formation
Proteolytic cleavage
Addition of carbohydrate/ phosphate/ lipids
Describe, the post translational modification involved in the production of insulin
Preproinsulin enters the ER: signal sequence is cleaved and 3 disulphide bonds form
This makes it proinsulin
The C chain is cleaved making it into insulin: A and B chain held together by disulphide bonds
