Abstract
The pressure effect on thermal inactivation, urea inactivation, guanidine hydrochloride (Gu·HCl) inactivation of Taka-amylase A (TAA) and also its pressure inactivation have been studied by measuring the rate of hydrolysis of p-nitrophenyl-α-maltoside (pNαM) catalyzed by TAA in the temperature range of 25-75°C and at the pressures up to 7kbar. The apparent rate of hydrolysis decreased with time above a certain temperature, pressure, and concentration of denaturants of urea and Gu·HCl. The decrease is due to the decrease in TAA activity. The enthalpy change of the thermal inactivation was 580-280kJ·mol-1, which corresponds to the denaturation process of N2 domain of TAA studied by thermal analysis. The volume change of this process was 120-79cm3·mol-1, while TAA was inactivated in the pressure range of 4-7kbar at 25-45°C and the corresponding volume change was -14--20cm3·mol-1. The pressure slightly depressed the urea inactivation and the volume change was 0-7cm3·mol-1. On the other hand, the pressure accelerated the Gu·HCl inactivation at lower concentration of Gu·HCl (1.0M) and the volume change was -10--20cm3·mol-1. At 2M Gu·HCl the volume change of inactivation was nearly zero.
