PRESSURE EFFECTS ON SUBSTRATE ACTIVATION PHENOMENA IN THE ALPHA-CHYMOTRYPSIN-CATALYZED HYDROLYSIS OF PARA-NITROPHENYL ACETATE

With and without p-chlorophenol as an activator, the rates of hydrolysis of p-nitrophenyl acetate catalyzed by .alpha.-chymotrypsin were measured at pressures up to 2 kbar at 25.degree. C. From the pressure dependence of the rate constant (kcat)A and (kcat)0 of the product formation with and without an activator, the activation volumes (.DELTA.Vcat)A and (.DELTA.cat)0 of the product formation with and without an activator, the activation volumes were +2 and -6 .+-. 1 cm3 .cntdot. mol-1. From the pressure dependence of the equilibrium constant (KA) of incorporation of p-chlorophenol into the enzyme, the volume change (.DELTA.VA) was -10 .+-. 1 cm3 .cntdot. mol-1. The mechanisms of the substrate activation are discussed in terms of the activation and reaction volumes.