Probing the limits of protein-amino acid side chain recognition with the aminoacyl-tRNA synthetases. Discrimination against phenylalanine by tyrosyl-tRNA synthetases

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Abstract
The specificity of the tyrosyl-tRNA synthetases from Escherichia coli and Bacillus stearothermophilus for tyrosine compared with phenylalanine was determined by using samples of phenylalanine which were scrupulously freed from tyrosine by either chemical or enzymic scavenging procedures. Both kinetic measurements and product analyses give a value of 1 .times. 105-2 .times. 105 for the preferential activation of tyrosine. Combined with the known ratio of phenylalanine to tyrosine in rapidly growing E. coli, an error rate of .apprx. 5/104 is calculated for the misactivation of phenylalanine. Since no evidence for an editing mechanism was found and this error rate is similar to observed rates in protein synthesis, the tyrosyl-tRNA synthetases appear to have adequate amino acid selection by simple preferential binding of the correct substrate. The incremental binding energy of the phenolic hydroxyl group of tyrosine is .apprx. 7 kcal/mol, a value presumed close to the maximum possible because of the evolutionary pressure on tyrosyl-tRNA synthetases for maximum specificity. A summary of high incremental binding energies determined from experiments on aminoacyl-tRNA synthetases is presented.