Valyl‐tRNA synthetase from yeast

Abstract

For discrimination between valine and the 19 naturally occurring noncognate amino acids, as well as between valine and 2-amino-isobutyric acid by valyl-tRNA synthetase from baker's yeast, discrimination factors (D) have been determined from k,,, and K , values in aminoacylation of tRNAVal-C-C-A. The lowest values were found for Trp, Ser, Cys, Lys, Met and Thr (D = 90 -870), showing that valine is 90 -870 times more frequently attached to tRNA""'-C-C-A than the noncognate amino acids at the same amino acid concentrations. The other amino acids exhibit D values between 1100 and 6200. Generally, valyl-tRNA synthetase is considerably less specific than isoleucyl-tRNA synthetase, but this may be partly compensated in the cell by valine concentrations higher than those of noncognate acids.In aminoacylation of tRNAVa'-C-C-A(3'NH2) discrimination factors D , are in the range of 40-1260. From D 1 values and AMP formation stoichiometry, pretransfer proof-reading factors n1 were determined; post-transfer proof-reading factors n2 were determined from D values and AMP formation stoichiometry in acylation of tRNAVal-C-C-A. n, values (7 -168) show that pretransfer proof-reading is the main correction step, posttransfer proof-reading (IT, z 1 -7) is less effective and in some cases negligible.Initial discrimination factors were calculated from discrimination and proof-reading factors according to a two-step binding process. These factors, due to different Gibbs free energies of binding can be related to hydrophobic interaction forces, and a hypothetical 'stopper' model of the amino-acid-binding site is discussed.In our recent work on amino acid specificity of aminoacyltRNA synthetases we found that no general error rate can be given for the aminoacylation of tRNA in the following reaction [ 1 -41 :

Review

“…The amino acid specificity of this class II synthetase was found to be poorer than those of some class I enzymes (Freist etal., 1987(Freist etal., ,1988(Freist etal., ,1989(Freist etal., ,1992Freist and Cramer, 1990). The amino acid specificity of this class II synthetase was found to be poorer than those of some class I enzymes (Freist etal., 1987(Freist etal., ,1988(Freist etal., ,1989(Freist etal., ,1992Freist and Cramer, 1990).…”