Prokaryotic translation

B. Prokaryotic translation

1. Initiation– three steps to initiation

(a) mRNA binds to 30S.

(i) Ribosome aligned by base pairing of a region of 16S rRNA of the 30S ribosomal subunit to a region on the mRNA 6-10 bases upstream of the initiation codon.

 (ii) The region is called the Shine/Dalgarno sequence.

(b) Methionyl-tRNA binds to 30S-mRNA complex.

(i) First aa-tRNA is positioned complementary to the first codon by the 30S ribosomal subunit.

(ii) In prokaryotes the first amino acyl-tRNA is always formyl methionyl tRNA (fMet-tRNAfMet)

(iii) Therefore, all proteins in prokaryotes are synthesized with formyl methionine as their first amino acid.

(iv) This complex is called the 30S pre-initiation complex.

(c) 50S subunit binds to 30S-tRNA-mRNA complex.

(i) This complex is called the initiation complex.

(ii) Two sites for amino acyl-tRNA binding on 50S subunit.

(a) The peptidyl site or P-site. This is the site where the growing polypeptide will be anchored to the ribosome. The polypeptide is held in place by covalent attachment of its carboxyl-terminal amino acid to its corresponding tRNA. This is called the peptidyl-tRNA.

(b) The amino acyl site or A-site. This is the site adjacent to the P-site where the next amino acyl-tRNA will bind at the ribosome. It is positioned directly over the triplet codon to allow codon-anticodon pairing.

(iii) At initiation, fMet-tRNA is in the P-site and the 2nd codon is positioned at the A-site.

2. Elongation

(a) Three reactions

(i) Binding of aa-tRNA to A-site

(a) Specificity determined by codon, anticodon pairing.

(b) GTP is hydrolyzed at the ribosome is pairing is correct.

(c) GTP hydrolysis is part of proofreading.

(d) If anti-codon and codon are completely non-complementary, amino acyl-tRNA will not bind to ribosome.

(e) If complement is sufficient, binding and GTP hydrolysis occur.

(f) If complement is not perfect, the ribosome will reject the amino acyltRNA.

(g) Hydrolysis of GTP must occur before the next step in translation can occur.

(ii) Transpeptidation

(a) Transfer of the growing polypeptide from peptidyl tRNA at the P-site

to the amino acyl-tRNA at the A-site.

(b) The ester bond between the peptide and tRNA of the peptidyl-tRNA is broken and the carboxyl-terminus of the peptide reacts with the amino group of the amino acid of the amino acyl-tRNA to form a peptide bond at the A-site. The hydrolysis of the ester bond provides sufficient energy to drive peptide bond formation.

(c) Catalyzed by peptidyl transferase.

 (iii) Translocation

(a) Movement of peptidyl-tRNA from A-site to the P-site and release of deacylated tRNA from the P-site.

(b) Also requires GTP hydrolysis.

(iv) Steps in elongation are repeated until a stop codon is reached.

(v) Many ribosomes at various stages of translation can be bound to a single mRNA.

(a) This structure is called a polysome.

(b) As a result many polypeptides are synthesized simultaneously from a single mRNA.

3. Termination

(a) When the ribosome encounters a UGA, UAG or UAA codon, no amino acid is added to the polypeptide. These codons are called termination codons.

(b) Termination also requires release factors.

(c) Function of release factors

(i) Catalyze hydrolysis of peptidyl tRNA

(ii) Promote dissociation of 50S subunit

(d) 30S dissociates or moves to the next start codon on the polycistronic mRNA


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