Evidence for single mechanism for aminoacyl-tRNA synthetases including aminoacyl adenylates as intermediates.

The rate of transfer of amino acid from enzyme-bound aminoacyl adenylate to tRNA has been compared with the rate of esterification of free amino acid. The approach of Lövgren et al. (Lövgren, T. N. E., Heinonen, J., and Loftfield, R. B. (1975) J. Biol. Chem. 250, 3854-3860) was used, with 14C in the aminoacyl adenylate and 3H in the free amino acid and with both the lysine and isoleucine systems of Escherichia coli. In both systems kinetic analyses show more rapid transfer from the preformed enzyme complex when interference by the back reaction with inorganic pyrophosphate was eliminated. Parallel experiments, in which the amount of enzyme complex was measured, confirmed that aminoacyl adenylate is an intermediate in both systems. No evidence was found for an alternative mechanism.