The process by which proteins are produced with amino acid sequences specified by the sequence of codons in messenger RNA is called translation. Translation is the first stage of protein biosynthesis.
In prokaryotes (unicellular), translation occurs in the cytosol, where the medium and small subunits of the ribosome bind to the tRNA. Translation process consists of following major phases that are discussed below;
Activation of amino acids:
The activation of aminoacids take place in cytosol.
The activation of aminoacids is catalyzed by their aminoacyl tRNA synthetases.
All the 20 aminoacids are activated and bound to 3’ end of their specific tRNA in the presence of ATP and Mg++.
The N-formylated methionine is chain initiating aminoacid in bacteria whereas methionine is chain initiating aminoacid in eukaryotes.
Methionine is activated by methionyl-tRNA synthetase. For N-formylmethionine two types of tRNA are used. Similarly, all 2o aminoacids are activated (amino acyl-AMP enzyme complex) and then bound to their specific tRNA forming Aminoacyl tRNA.
In the first step, initiation factor-3 (IF-3) binds to 30S ribosomal unit.
Then mRNA binds to 30S ribosomal subunit in such a way that AUG codon lie on the peptidyl (P) site and the second codon lies on aminoacyl (A) site.
The tRNA carrying formylated methionine FMet–tRNA is palced at P-site. This specificity is induced by IF-2 with utilization of GTP.
Shine Dalgarno sequence in the mRNA guide correct positioning of AUG codon at P-site of 30S ribosome.
Binding of AA-tRNA at A-site:
The 2nd tRNA carrying next amino acid comes into A-site and recognizes the codon on mRNA. This binding is facilitated by EF-TU and utilizes GTP.
After binding, GTP is hydrolyzed and EF-TU-GDP is released
EF=TU-GDP then and enter into EF-TS cycle.
ii. Peptide bond formation:
The amino acid present in t-RNA of P-site ie met is transferred to t-RNA of A-site forming peptide bond. This reaction is catalyzed by peptidyltransferase.
Now, the t-RNA at P-site become uncharged.
iii. Ribosome translocation:
After peptide bond formation ribosome moves one codon ahead along 5’-3’ direction on mRNA, so that dipeptide-tRNA appears on P-site and next codon appear on A-site.
The uncharged tRNA exit from ribosome and enter to cytosol.
The ribosomal translocation requires EF-G-GTP (translocase enzyme) which changes the 3D structure of ribosome and catalyze 5’-3’ movement.
The codon on A-site is now recognized by other aminoacyl-tRNA as in previous.
The dipeptide on P-site is transferred to A-site forming tripeptide.
This process continues giving long polypeptide chain of aminoacids.
The peptide bond formation and elongation of polypeptide continues until stop codon appear on A-site.
If stop codon appear on A-site it is not recognized by t-RNA carrying amino acids because stop codon do not have anticodon on mRNA.
The stop codon are recognized by next protein called release factor which hydrolyzes and cause release of all component such as 30s, 50S, mRNA and polypeptide separates.
RF-1 recognizes UAA and UAg while RF-2 recognizes UAA and UGA while RF-3 dissociate 30S and 50S subunits.