protein translation Flashcards
Describe the structure of typical mRNA.
mRNA has a 7-methylguanylate cap at the 5’ end followed by a 5’ untranslated region (UTR) You then get the coding region in the middle At the end you get a 3’ UTR and the poly-A tail
What are the stop codons?
UAA, UGA, UAG
What is the Methionine/start codon?
AUG
What is the significance of Methionine?
It is the first amino acid in virtually all polypeptides
What enzyme is involved in the transfer of an amino acid to tRNA?
Aminoacyl tRNA synthetase.
the correct amino acid gets stuck using aminoacyl tRNA synthetases.
there is 1 aminoacyl tRNA synthetases.for each amino acid
Describe the structure of tRNA
-clover shaped
-carries amino acid on 3’ end
-64 tRNAs, one for each codon so an amino acid can have several tRNA
when it holds an amino acid it is called Charged
How does aminoacyl tRNA synthetases work?
Free amino acid binds to AMINOACYL tRNA SYNTHASE
Aminoacyl tRNA synthase cleaves pyrophosphate from ATP and binds remaining AMP to amino acid
Amino acid becomes ADENYLATED
Adenylated amino acid is then attached to tRNA
Aminoacyl tRNA SYNTHASE and AMP then detach, leaving the charged tRNA
What are the three stages of translation?
Initiation – Elongation - Termination
what is site called that Met-tRNA binds to
P site
p site is on the left and A site is on the right
Describe initiation.
- elF4 E and elF4G bind to the 7-MeG cap and this is recognised by the preinitation complex.
- Preninitation complex move along until Met-tRNA pairs with first AUG.
- Bind, then GTP on elF2 is hydrolysed to GDP + pi , which provides energy to ensure correct base pairing
- when GTP is hydrolysed there is a conformational change which allows the 60s subunit to bind.
- the elF2 with GDP dissociates
What components are make up the preinitiation complex?
40S ribosomal subunit
Methionine-tRNA eIF2 (initiation factor) with GTP.
Describe elongation
The next charged tRNA comes and binds to the A site of ribosome
Peptidyl transferase catalyses the formation of a peptide bond between the two amino acids
The tRNA from the P site then dissociates and the ribosome moves along, this is called translocation and uses elongation factors which use GTP
What is the benefit of using GTP
ELONGATION FACTORS then move the ribosome along mRNA using GTP, called TRANSLOCATION
In the time taken for GTP to hydrolyse, there is opportunity for incorrect base pairs to dissociate
So, the pauses given by GTP hydrolysis increases accuracy of translation
Describe termination
Stop codon attracts RELEASE FACTORS
Bind to empty A site on ribosome
Peptidyl Transferase bonds water to final amino acid, creating carboxyl group
Translation complex dissociates into cytoplasm
Which proteins have a signal sequence
Proteins that are destined for secretory or transmembrane
Where is the signal sequence found and what does it consist of?
It is found at the N terminus.
It mainly consists of hydrophobic amino acids
-first 20-24 amino acids are a signal recognition particle, containing the signal sequence
Describe how proteins enter the ER.
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 ER 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 ER.
Step 4: cleavage of signal sequence and protein folding
What extra features do proteins that are destined to be transmembrane have?
They have an extra hydrophobic sequence to hold them in the membrane.
what happens to the signal sequence after the proteins is inside the rer
it is degraded by signal peptidase
Describe, in full, the post translational modification involved in the production of insulin.
- Begin with preproinsulin, which has an N-terminus signal sequence so signal sequence is removed and degraded once inside the ER
- 3 disulphide bonds form between cysteine residues while the protein folds to form PROINSULIN
- Then the C chain is cleaved making it into insulin. You get an A and B chain held together by 3 disulphide bonds
whats an easy way to measure insulin
Chain C is released into cytoplasm and then into blood as a waste product
It is difficult to detect insulin levels in blood
So instead, we can
measure Chain C levels as an indirect measure of insulin
what are antibiotics?
Antibiotics are natural products of bacteria or fungi that can selectively inhibit prokaryotic protein synthesis because the translational machinery is complex and easily disrupted.
what are the post translational modification of insulin?
- Signal sequence is removed and degraded
- Disulphide bonds form between cysteine residues within the polypeptide chain- 3 disulphide bonds form while the protein folds to form pro-insulin ( single chain polypeptide with 3 disulphide bonds)
- Before it is packaged into secretory vesicles, it is proteolytically cleaved in 2 positions to release the C chain
- This leaves you with active, fully functional insulin, where the A and B chains are held together by 3 disulphide bonds.
where does protein synthesis take place and what is the relevance of this?
Protein synthesis takes place in the cytoplasm
But, cellular organelles are bound by a membrane so the cell needs a mechanism to transfer proteins across membranes
where are secretory and transmemebrane proteins synthesised?
RER
what are the examples of antibiotic and what it inhibits?
Streptomycin Inhibits initiation
Tetracycline Inhibits aa-tRNA
binding
Erythromycin Inhibits
translocation
Chloramphenicol Inhibits peptidyl
transferase
Puromycin Terminates elongation
prematurely
