Methionyl-tRNA synthetase from Escherichia coli: primary structure at the binding site for the 3'-end of tRNAfMet

Abstract

It was previously shown that when the tryptic fragment of methionyl-tRNA synthetase from E. coli is incubated with periodate-treated initiator tRNA, it is inactivated due to the formation of a covalent 1:1 complex that could be stabilized by reduction with cyanoborohydride. In this work, the residues labeled in the trypsin-modified enzyme were identified. After chymotryptic digestion of the protein-tRNA complex, 2 major labeled peptides (A and B) and a minor one (C) were isolated and identified by sequencing. The radioactivity associated with peptides A-C represented 65-75, 20-25, and 2-4%, respectively, of the radioactivity eluted from the peptide maps. Peptides A and B encompassed lysines-335 and -61, respectively. Both these lysines were fully labeled. Peptide C encompassed lysines-142, -147, and -149 each of which was incompletely labeled. The significance of these results is discussed in light of the known crystallographic structure of the enzyme.