Covalent complex of phenylalanyl-tRNA synthetase with 4-thiouridine-substituted tRNA Phe gene transcript retains aminoacylation activity

Abstract

s 4U-containing transcripts of tRNA Phe gene have been prepared by complete substitution of 16 U residues or by random incorporation of s 4U residues followed by affinity electrophoresis isolation of s 4U-monosubstituted tRNA transcripts. Both analogs have been cross-linked to Thermus thermophilus phenylalanyl-tRNA synthetase (PheRS) and the specificity of the cross-linking has been demonstrated. Functional activity of the covalent complex of PheRS with the s 4U-monosubstituted transcript has been shown by aminoacylation of 60% of the enzyme-cross-linked tRNA. This is the first instance in which biological activity of aminoacyl-tRNA synthetase and cross-linked tRNA in a specific complex has been revealed.