Arginyl‐tRNA synthetase from yeast

For discrimination between arginine and 19 other amino acids in aminoacylation of tRNA Arg ‐C‐C‐A by arginyl‐tRNA synthetase from baker's yeast, discrimination factors ( D ) have been determined from k cat and K m values. The lowest values were found for Trp, Cys, Lys ( D = 800–8500), showing that arginine is 800–8500 times more often incorporated into tRNA Arg ‐C‐C‐A than noncognate acids at the same amino acid concentrations. The other noncognate amino acids exhibit D values between 10000 and 60000. In aminoacylation of tRNA Arg ‐C‐C‐A(3′NH 2 ) discrimination factors D 1 are in the range 10–600. From these values and AMP formation stoichiometry, pretransfer proof‐reading factors II 1 were determined; from D values and AMP stoichiometry in aminoacylation of tRNA Arg ‐C‐C‐A, posttransfer proof‐reading factors II 2 could be calculated. II 1 values between 2 and 120 show that pretransfer proof‐reading is the main correction step, posttransfer proof‐reading ( II 2 ∼ 1–10) plays a marginal role. Initial discrimination factors due to different Gibbs free energies of binding between arginine and the noncognate amino acids were calculated from discrimination and proof‐reading factors. According to a two‐step binding process, two factors ( I 1 and I 2 ) were determined. They can be related to hydrophobic interaction forces and hydrogen bonds that are especially formed by the arginine side chain. A hypothetical ‘stopper’ model of the amino acid recognition site is discussed.