EDITING MECHANISM FOR THE METHIONYL-TRANSFER RNA-SYNTHETASE IN THE SELECTION OF AMINO-ACIDS IN PROTEIN-SYNTHESIS

Trace impurities of methionine in commercial samples of amino acids have obscured the pattern of misactivations catalyzed by methionyl-tRNA synthetases and have concealed the existence of an editing mechanism. Recrystallized samples of α-aminobutyrate, valine, and isoleucine, for example, are found to stimulate the exchange of pyrophosphate into ATP catalyzed by the enzyme from Bacillus stearothermophilus with values of Kcat approaching that for the methionine-stimulated reaction. Following removal of methionine from the nonspecific amino acids by treatment with Raney nickel, activity is abolished with the branched side chain amino acids and considerably reduced with certain unbranched side chain amino acids. There is an editing mechanism for the removal of the misactivation of amino acids with smaller side chains than methionine: the enzymes from Escherichia coli and B. stearothermophilus do not catalyze the transfer of homocysteine or norvaline to tRNAMet but instead catalyze an ATP-pyrophosphatase activity. The turnover number for the ATP-pyrophosphatase activity in the presence of homocysteine and tRNAMet is similar to that for the aminoacylation of the tRNA with methionine. The amino acids are sorted by size in accord with the “double-sieve” editing mechanism. Amino acids sterically smaller than methionine are activated with reduced values of kcat/KM and their products are removed by hydrolytic editing. The sterically larger ethionine is activated, and its products are resistant to hydrolysis. Norleucine, which although nominally isosteric with methionine is slightly smaller, is on the borderline of being edited: there is both transfer to tRNAMet and an amino acid stimulated ATP/pyrophosphatase activity. The nonaccepting terminal hydroxyl group of tRNAMet is not essential for the editing reaction. The synthetic tRNAMet which is deoxy in this position does not accept homocysteine or norvaline but stimulates the amino acid dependent ATP-pyrophosphatase reaction. © 1979, American Chemical Society. All rights reserved.