Discrimination between D- and L-tyrosyl transfer ribonucleic acids in peptide chain elongation

Abstract

D-Tyr-tRNA can take part in peptide bond formation with N-AcPhe-tRNA on ribosomes programmed with the hexanucleotide UUUUAC. None of the steps leading to peptide bond formation exhibit high stereoselectivity. Ternary complex formation with EF-Tu.GTP favors L-Tyr-tRNA by a factor greater than 25. The complex formed with D-Tyr-tRNA was not protected from hydrolysis, which suggests that the D-amino acid is improperly bound to the protein. The rate of EF-Tu-promoted dipeptide formation was 30-fold faster with L-Tyr-tRNA. The ratio of moles of GTP hydrolyzed to dipeptide formed was 1.4 for L-Tyr-tRNA and 4 for D-Tyr-tRNA. The excess of GTP hydrolyzed to peptide bonds formed is evidence for kinetic proofreading in AA-tRNA selection. The combined effects of the partial discrimination at each stage, from the aminoacylation to the peptide formation, favor L-tyrosine by a factor greater than 10(4) and would virtually exclude D-tyrosine from being incorporated under conditions where L-tyrosine was also present.