Effect of enzyme-substrate interactions away from the reaction site on carboxypeptidase A catalysis

The kinetics of 14 peptide substrates of carboxypeptidase A have been studied for the purpose of evaluating P-1-P-3/S-1-S-3 interactions. It was found that the amide group at P-1-P-2 is required for efficient catalysis. This observation is consistent with previously proposed hydrogen bonding interactions, based on crystallographic data, between the P-1 NH and Tyr-248 and between the P-2 carbonyl oxygen and Arg-71. In contrast, substitution of the benzamido amide group (at P-2-P-3) Of N-benzoylglycylglycyl-1-phenylalanine by -CH2CH2- resulted in more effective catalysis. In this case hydrophobic interactions are important in the ground state and in the transition state of the rate-determining step. (C) 1996 Academic Press. Inc.