Effect of high hydrostatic pressure on D-amino acid oxidase activity and stability

We report the effects of high hydrostatic pressure (HHP) on D-amino acid oxidase (DAAO) from pork kidney stability and activity. At 20 °C, the activity decreased with increasing pressure, but at 40 °C, it was not significantly affected because the increased catalytic rate compensated for the enzyme inactivation within the time frame of the experiment. At 50-300 MPa, raising the temperature increased the rate of DAAO inactivation, indicating that increases in both pressure and temperature contributed to inactivation. In contrast to other oxidases pressure did not stabilize DAAO against thermal inactivation. Treatments between 0.1 and 100 MPa had no significant effect on its stability at 55 °C. However, the rate of inactivation increased with increasing pressure from 100 to 300 MPa, resulting in an activation volume (ΔV^‡) of inactivation of -15.8 ± 1.7 cm³mol⁻¹. Unlike other oxidases that have buried cavities larger than 100 ų (0.1 nm³), DAAO has cavities smaller than 75 ų (0.075 nm³), thus supporting the hypothesis that cavity size determines whether pressure stabilizes an enzyme against thermal inactivation.