Peptidyl-tRNA

The a-amino group of the new aminoacyl-tRNA in the A site carries out a nucleophilic attack on the esterified carboxyl group of the peptidyl-tRNA occupying the P site (peptidyl or polypeptide site). At initiation, this site is occupied by aminoacyl-tRNA mef. This reaction is catalyzed by a peptidyltransferase, a component of the 285 RNA of the 605 ribosomal subunit. This is another example of ribozyme activity and indicates an important—and previously unsuspected—direct role for RNA in protein synthesis (Table 38-3). Because the amino acid on the aminoacyl-tRNA is already activated, no further energy source is required for this reaction. The reaction results in attachment of the growing peptide chain to the tRNA in the A site. 

The now deacylated tRNA is attached by its anticodon to the P site at one end and by the open GGA tail to an exit (E) site on the large ribosomal subunit (Figure 38-8). At this point, elongation factor 2 (EE2) binds to and displaces the peptidyl tRNA from the A site to the P site. In turn, the deacylated tRNA is on the E site, from which it leaves the ribosome. The EF2-GTP complex is hydrolyzed to EF2-GDP, effectively moving the mRNA forward by one codon and leaving the A site open for occupancy by another ternary complex of amino acid tRNA-EFlA-GTP and another cycle of elongation. 

Figure 38-8. Diagrammatic representation of the peptide elongation process of protein synthesis. The small circles labeled n - 1, n, n -I-1, etc, represent the amino acid residues of the newly formed protein molecule. EFIA and EF2 represent elongation factors 1 and 2, respectively. The peptidyl-tRNA and aminoacyl-tRNA sites on the ribosome are represented by P site and A site, respectively.

Figure 38-9. Diagrammatic representation of the termination process of protein synthesis. The peptidyl-tRNAand aminoacyl-tRNA sites are indicated as P site and A site, respectively. The termination (stop) codon is indicated by the three vertical bars. Releasing factor RF1 binds to the stop codon. Releasing factor RF3, with bound GTP, binds to RFl. Flydrolysisofthe peptidyl-tRNA complex is shown by the entry of HjO. N and C indicate the amino and carboxyl terminal amino acids, respectively, and illustrate the polarity of protein synthesis.

For processive peptide polymerization, the rihosome has to move along the mRNA. Following peptide bond formation, the rihosomal A site is occupied hy a peptidyl-tRNA whereas the P site contains a deacylated tRNA. During translocation, the complex of the two tRNAs with the mRNA has to move relative to the ribosome to... 

In the translocation step, the ribosome moves exactly three nudeotides (one codon) along the message. This moves the growing peptidyl-tRNA into the P site and aligns the next codon to be translated with the empty A site. 

A nasopharyngeal swab obtained from a 4-month-old infrint with rhinitis and paroxysmal coughing tested positive upon culture for Bordetella pertussis. He was admitted to the hospital for therapy with an antibiotic that inhibits the translocation of peptidyl-tRNA on 70S ribosomes, This patient was most likely treated with... 

Fig. 11. Comparison of the peptidyl transfer reaction in the ribosome and in the selected peptidyltransferase ribozyme. The ribosome contains a binding site for the peptidyl-tRNA (P-site) and for the aminoacyl-tRNA (A-site). In the selected ribozyme the binding site for the AMP-Met-Bio substrate would be analogous to the P-site. The attacking a-amino group which is bound in the A-site in the ribosome is covalently attached to the 5 -end in the ribozyme. Catalytically active RNAs preferentially attach the biotin tag onto themselves and can thus be separated from the inactive ones...

The final step in protein biosynthesis is chain termination. Natural mRNA molecules contain termination codons UAA, UGA, or UAG There are no tRNAs that have anticodons which are complementary to these codons. When the growing peptide chain encounters one of these termination codons, the peptidyl-tRNAis transferred to water instead of another aminoacyl-tRNA. The peptidyl-tRNA is hydrolyzed to free the completed protein and the tRNA. Chain termination completes protein synthesis. 

Then the peptidyl-tRNA at the A site is translocated to the P site by the ribosome moving along the mRNA a codon at a time, exposing the A site for a new aminoacyl-tRNA appropriate for the particular codon, and a repeat of the elongation process occurs. The cycles of elongation and translocation continue until a termination codon is reached, and the peptide or protein is then hydrolysed and released... 

Macrolides, lincosamides and streptogramins are protein biosynthesis inhibitors that bind to 50S subunit of the ribosome and inhibit peptidyl tRNA translocation from the A-site to the P-site." Macrolides have a glycosylated 14-, 15- or 16-membered lactone ring structure and are produced by several species of Streptomyces. Lincosamide antibiotics were isolated initially from Streptomyces lincolnensis but later isolated from different species of Streptomcyces. Streptogramins were also isolated from Streptomycesgraminofaciens and subsequently from several different Streptomyces species. There are two structurally different streptogramins, A and B they are bacteriostatic individually and can be bactericidal when combined. 

This enzyme [EC 3.1.1.29], also called peptidyl-tRNA hydrolase, catalyzes the hydrolysis of an A-substituted aminoacyl-tRNA to produce an A-substituted amino acid and tRNA. 

This ribosomal enzyme [EC 2.3.2.12] catalyzes the acyl transfer from a peptidyl-tRNA with an aminoacyl-tRNA to produce tRNA and a peptidylaminoacyl-tRNA. ... 

BOROHYDRIDE REDUCTION ACYL-SERINE INTERMEDIATE ACTIVE SITE TITRATION Acyl transfer from a peptidyl-tRNA,... 

The nascent chain attached to the previous tRNA used, peptidyl tRNA, is transferred to the P site of the ribosome. 

The ribosome then translocates to the next codon, with the peptidyl-tRNA shifting from the A site to the P site and the now uncharged tRNA exiting the ribosome from the E site. 

Frolova LY, Tsivkovskii RY, Sivolobova GF, Oparina NY, Serpinsky Ol, Blinov VM, Tatkov SI, Kisselev LL (1999) Mutations in the highly conserved GGQ motif of class 1 polypeptide release factors abolish ability of human eRFl to trigger peptidyl-tRNA hydrolysis. RNA 5 1014-1020... 

Karimi R, Ehrenberg M (1994) Dissociation rate of cognate peptidyl-tRNA from the A-site of hyper-accurate and error-prone ribosomes. Eur J Biochem 226 355-360 Karimi R, Ehrenberg M (1996) Dissociation rates of peptidyl-tRNA from the P-site of E.coli ribosomes. EMBOJ 15 1149-1154... 

Song H, Mugnier P, Das AK, Webb HM, Evans DR, Tuite ME, Hemmings BA, Barford D (2000) The crystal structure of human eukaryotic release factor eRFl—mechanism of stop codon recognition and peptidyl-tRNA hydrolysis. Cell 100 311-321... 

A change in the three-dimensional conformation of the entire ribosome results in its movement along the mRNA. Because the structure of EF-G mimics the structure of the EF-Tu-tRNA complex (Fig. 27—25b), EF-G can bind the A site and presumably displace the peptidyl-tRNA. 

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