A specific modification next to the anticodon of phenylalanine transfer ribonucleic acid.

Abstract

By mild treatment of tRNAPheyeast with acid (pH 2.9, 37°, 3–4 hours) the base Y+, which according to RajBhandary, Khorana et al. is located right next to the anticodon, can be excised without breaking the chain. Y+ and the acid treated tRNAPhe (=tRNAPheHCl) were characterized and the conversion of tRNAPhe to tRNAPheHCl was investigated in detail. 1 Y+ is a rather hydrophobic compound of unknown structure. Y+ and ribose were found when the nucleoside Y was treated with acid. Some physical and chemical properties of Y+ are reported. Y+ can be isolated easily in pure form from oligonucleotides, tRNAPheYeast, or even unfractionated tRNAyeast (not tRNAcoli) by mild acid treatment and subsequent extraction into chloroform. 2 tRNAPheHCl and tRNAPhe have very similar molecular sizes and shapes since their Sephadex elution profiles were overlapping. The facile separation of tRNAPhe and tRNAPheHCl by counter current distribution, on columns of methylated albumin on Kieselgur or benzoylated DEAE-celluose is due, at least in part, to the removal of a hydrophobic residue from an exposed region of the tRNAPhe. 3 tRNAPheHCl can be charged with Phe to the same extent as tRNAPhe. Since also the Michaelis constant of phenylalanyl tRNA synthetase is the same for both tRNAs (1.5×10−7 M) and the maximal velocity of charging is decreased by only one third in tRNAPheHCl, it can be concluded, that Y and probably the whole anticodon loop are not essential parts of the synthetase recognition region of tRNAPhe in homologous charging reactions. 4 It was found that tRNAPheHCl and tRNAPhe from which the –CpCpA endgroups had been removed do not differ in their acceptor capacities for CTP and ATP. 5 In the ribosome systems, Phe-tRNAPheHCl, does not bind to poly (U) or other polynucleotides and does not transfer its amino acid onto growing peptide chains. The complete loss of codon recognition is attributed largely to a change in the conformation of the anticodon loop. The ultraviolet spectra of an Y-containing hexanucleotide and of its acid conversion products are in agreement with the notion of a strong interaction between Y+ and its neighbouring bases, as are the rate differences in the liberation of Y+ from Y [1.5 times faster] and the hexanucleotide, respectively. 6 The rate of tRNAPheHCl formation, as measured by methylated albumin on Kieselgur-column chromatography and binding assays, is the same as the rate of Y+-liberation from the hexanucleotide. The acid induced conversion of tRNAPhe to tRNAPheHCl is a completely selective reaction affecting Y as the only nucleoside of tRNAPhe.