Phenylalanyl transfer ribonucleic acid synthetase from rat liver. Analysis of phenylalanine and adenosine 5'-triphosphate binding sites and comparison to the enzyme from Escherichia coli.

Inhibition of the ATP--PPi exchange reaction catalyzed by rat liver phenylalanyl-tRNA synthetase by structural analogues of L-phenylalanine and ATP has been examined and compared with data reported for the enzyme from E. coli. The phenylalanine binding sites are similar in the following characteristics. (1) The region that complexes the phenyl ring shows a strict requirement for the unsubstituted phenyl ring altough the rat liver enzyme is more tolerant in this respect. (2) The protonated amino group of phenylalanine is required for binding. (3) The region neighboring the binding site for the carboxylate of phenylalanine is diffusely hydrophobic. Unlike effects of these modifications on interaction with the E. coli enzyme, substitution of the carboxylate by hydrophobic groups leads to large losses in affinity for the rat liver enzyme. Although both enzymes bind D-phenylalanine very poorly, their relative affinities for the D isomers of phenylalanine analogues vary greatly. The most dramatic difference is observed with N-benzyl-D-amphetamine, which binds 24 000-fold tighter to E. coli phenylalanyl-tRNA synthetase than the rat liver enzyme. The affinity of rat liver phenylalanyl-tRNA synthetase for naturally occurring adenine compounds is similar to that of the E. coli enzyme, suggesting that the binding of ATP occurs via similar interactions. Adenine provides a major protion of the free energy of binding of ATP. The remainder may be viewed as the sum of detrimental interactions with the phosphate groups of ATP and a favorable contribution by the ribose moiety.