Trypsin hydrolysis of X-phenyl hippurates

The kinetics of hydrolysis of eight substituted phenyl hippurates by trypsin at pH values 6, 7 and 8 were investigated and the kinetic constants Km, kcat (kcatalysis) and kcat/Km were shown to fit the Hammett equation. The rho (rho) value obtained from the correlation of trypsin binding with this class of esters was compared with that obtained with other serine and cysteine proteases. The rho value for trypsin was similar to that obtained for alpha-chymotrypsin in that both enzymes reveal a pronounced dependence on through resonance (sigma-) in the formation of the Michaelis complex and the acyl-enzyme. It is apparent that through resonance facilitates the leaving of the phenoxy moiety during catalysis in the serine proteases but not in the case of the cysteine or bacterial serine proteases.