The process of protein synthesis from a mRNA molecule is known as translation. It occurs mainly in the ribosomes. Consider the messenger RNA structure like a tape. As soon it attaches to the ribosome, the process of protein synthesis starts. The movement of the RNA molecule starts creating a lengthy polypeptide chain. First, the leading end on mRNA (the 5' end) emerges from the first ribosome. Next, it attaches to the second ribosome for synthesizing the next polypeptide chain. Once the RNA moves from a ribosome, it gets vacated for the new set of instructions. The amino acids start getting assembled into a polypeptide chain from the amino terminus. The process of assembly finishes at the carboxylate terminus. During the process of translation, an RNA molecule passes on the amino acid molecule to the growing polypeptide chain. Hence, it is known as a tRNA molecule. The transfer RNAs belong to a class of smallest biologically active molecules. The transfer RNAs attach to the amino acids at their 3' end.

The translation of mRNA occurs from 5’ to 3’ direction. An amino acid binds to its specific tRNA. The codon of mRNA binds to the anticodon of tRNA through a complementary base pairing. The mRNA is specific in recognizing the anticodon. An enzyme known as aminoacyl tRNA synthetase attaches a correct amino acid to the tRNA. This process is known as aminoacylation or charging. Let us consider the example of valine tRNA. The first step involves binding of amino acid and ATP to a specific aminoacyl tRNA synthetase. Hence, the reaction leads to the loss of two phosphate groups from ATP. It leads to the formation of aminoacyl AMP. The next step involves binding of uncharged tRNA to the enzyme. As a result, the enzyme transfers the amino acid to tRNA. Hence, the aminoacyl tRNA releases from the enzyme. A factor known as transfer factor I participates in binding the charged tRNA to the ribosome. The transfer factor II is also known as translocase. It is a protein capable of forming a complex with the GTP and the ribosome. When the charged tRNA gets translocated from its ribosomal entrance site to the peptidyl site, it leads to the hydrolysis of GTP to GDP, thereby releasing the translocase.

Image 1: Translation initiation and peptide bond formation: (1) The first image depicts the binding of the fMet tRNA to the initiation site. (2) The serine tRNA binds in the A site. (3) Peptide bond formation (4) The uncharged tRNA moves to the E site and the tRNA consisting of amino acids linked through peptide bond moves to the P site.





Initiation of translation:

The mRNA with an AUG initiation codon gets involved in the process of translation. The mRNA ribosome binding site (also known as Shine Dalgarno sequence) also contributes to the process of translation. The 30S ribosomal subunit binds to AUG start codon. Later on, the initiator tRNA binds to this site. Since, AUG start codon codes for methionine amino acid, the newly made proteins start with amino acid methionine. In prokaryotes, the methionine gets modified to formylmethionine (fMet). The fMet tRNA has a 5’-CAU-3’ anticodon. Binding of fMet tRNA to the start codon releases an IF3. Hence, it forms a 30S initiation complex. It consists of mRNA, 30S subunit, fMet tRNA, IF1, and IF2. The next step involves binding of the 50S ribosomal subunit, leading to the hydrolysis of GTP. This reaction releases the IF1 and IF2 factors. It finally leads to the formation of the 70S initiation complex. A site-P site hypothesis describes the ribosomal sites. Following are the three main binding sites for the aminoacyl tRNA:

· The exit site is also known as the E site.

· The peptidyl site is also known as the P site.

· Aminoacyl site is also known as A site.

The fMet tRNA binds to the mRNA at the P site. A site accepts the incoming aminoacyl tRNA. For transferring the peptide group, it requires translocation of tRNA. Hence, the tRNA gets translocated from the A site to the P site.





Elongation:

The amino acids get added to the polypeptide chain thereby allowing it to grow till the required amount. It involves three main steps. Primarily, the aminoacyl tRNA comes in contact with the ribosome and binds to it. Secondly, it leads to the formation of the peptide bond through an enzymatic reaction. The third step involves the movement of the ribosome (translocation) along the mRNA. It considers one codon at a time . The peptidyl site of the ribosome comes in contact with the AUG codon. It is suitable for the fMet tRNA to bind to the mRNA. The tRNA anticodon helps in binding to the mRNA codon. Next step involves binding of the aminoacyl tRNA (for example ser tRNA) in the A site. Binding of an aminoacyl tRNA to the codon in the A site releases the elongation factor known as EF-Tu. The reaction involves the hydrolysis of GTP. Note that the elongation factors EF-Tu get recycled for the next aminoacyl tRNA. Now, the fMet tRNA and aminoacyl tRNA come close to each other. Since these two amino acids are adjacent to each other, a peptide bond forms between the two amino acids. In this case, a peptide bond forms between the formylmethionine and serine amino acids. The reactions get catalyzed by peptidyl transferase.

The amino acids linked by a peptide bond get attached to the aminoacyl tRNA situated at the A site. The tRNA is now known as peptidyl tRNA. Next step involves the process of translocation. The ribosome moves from one codon to another codon. This step again involves EF-G factors and GTP. Hence, the peptidyl tRNA moves from A site to the P site, leaving the A site empty. Similarly, the uncharged tRNA moves to the E site from the P site. An uncharged tRNA is a tRNA that has given its amino acid for peptide chain elongation.

The tRNA without an attached amino acid gets released. Next, the ribosome starts preparing for the next elongation cycle. The empty A site gets occupied by another aminoacyl tRNA with a specific anticodon. The above process gets repeated till the sufficient proteins get synthesized.

Image 2: Elongation and termination of translation: (5) Next aminoacyl tRNA binds to the A site. (6)Peptide chain elongation. (7) Termination of the process due to the activity of the release factor. (8) Dissociation.





Termination:

None of the tRNAs possess anticodon for a stop codon. A protein group known as termination factor or release factor (RF) help the ribosome in recognizing a stop codon. There are three types of release factors in E. coli such as RF1, RF2, and RF3. Each RF is a single polypeptide. The role of RF1 involves recognition of UAA and UAG codons. The RF2 involves recognition of UAA and UGA. The RF3 does not recognize any stop codon. The ribosome recognizes a chain termination codon (UAG). Then the polypeptide chain present on the peptidyl tRNA (present on the P site) gets released. The ribosomal subunits get dissociated thereby separating the remaining components.





References:

[1] Principles of genetics, Gardner, M. J. Simmons, D. P. Snustad, eighth edition.

[2] Biology of the Prokaryotes, edited by Joseph W. Lengeler, Gerhart Drews.

[3] Genetics, G. Archunan

[4] Genetics, Daniel Hartl, Maryellen Ruvolo



