Phenylalanyl‐tRNA Synthetase from Yeast and Its Discrimination of 19 Amino Acids in Aminoacylation of tRNA Phe ‐C‐C‐A and tRNA Phe ‐C‐C‐A(3′NH 2 )

For discrimination between phenylalanine and 18 other naturally occurring non‐cognate amino acids by the class II aminoacyl‐tRNA synthetase specific for phenylalanine, discrimination factors, D , of 190–6300 have been determined from k cat and K m values. Generally, phenylalanyl‐tRNA synthetase is more specific than the class II enzymes specific for Lys and Thr, but works with lower accuracy than the class I enzymes specific for IIe, Tyr, and Arg. In aminoacylation of tRNA Phe ‐C‐C‐A(3′NH 2 ) discrimination factors D 1 vary between 80–1610. Pre‐transfer proof‐reading factors II 1 are in the range 2.3–74, post‐transfer proof‐reading factors II 2 in the range 1.0–4.6, showing that pre‐transfer proof‐reading is the main correction step, post‐transfer proof‐reading is less effective or negligible. Initial discrimination factors ( I 1 and I 2 ) caused by differences in Gibbs free energies of binding between phenylalanine and non‐cognate amino acids have been calculated assuming a two‐step binding process. Factors I 1 can be related to hydrophobic‐interaction forces depending on accessible surface areas of the amino acids, factors I 2 scatter about a low mean value and do not show any relation to amino acid structures or surfaces, indicating less checking of amino acid side chains in the putative second binding step.