[Study of the possibility of identifying the structural elements of the phenylalanyl-tRNA-synthetase active center by affinity labeling].

The possibility of localization of active sites structural components by affinity labelling was investigated. The modification of E. coli MRE-600 phenylalanyl-tRNA synthetase (E.C.6.1.1.20) (alpha 2 beta 2-type) by the phosphorylating analog of ATP-- [14C]adenosine-5'-trimetaphosphate results in the labelling of both heavy (beta) and light (alpha) enzyme subunits. Analysis of the peptide maps of the tryptic enzyme hydrolysate reveals a great number of peptides containing [14C]radioactivity. The decrease of covalent binding at low concentration of the analog did not abolish the plural labelling. The data permit to consider this kind of analogs as unperspective for localization of specific peptides. Modification of phenylalanyl-tRNA synthetase by tRNAPhe containing the photoreactive group (--CH2CONHC6H5N3) at eighth position of molecule (S8U) results in the labelling of only heavy beta-subunits. These data correspond to the previous results which testify to the disposition of tRNA binding sites on beta-subunits of phenylalanyl-tRNA synthetase. After hydrolysis of the modified phenylalanyl-tRNA synthetase by trypsin six peptides covalently bound with tRNAPhe were revealed. This quantity of modified peptides is higher than the number of tRNA binding sites. Hence the method of affinity labelling has definite limitations for localization of peptides of enzyme active sites.