Pressure effect on subtilisin crystallization and solubility

High pressures (135 and 200 MPa) arrested the crystallization process in un seeded batch crystallizations of subtilisin conducted at 25 degrees C over a period of seven days. Crystallization resumed at the normal atmospheric p ressure rate once batches were depressurized and the active soluble enzyme concentration decayed to the same value as seen in atmospheric crystallizat ions. Solubility experiments conducted at 25 degrees C showed an exponentia l increase in apparent solubility with pressure. The solubility increased a pproximately five-fold at 200 MPa. The partial molar volume change of cryst allization for this increase in solubility with pressure is + 21 +/- 1 ml/m ol. Solubility changes with pressure are largely responsible for the arrest in crystallization rate at high pressure. Experiments on pH-pressure and i onic strength-pressure interactions showed that the change in apparent solu bility is not due to pressure-induced changes in pH and ionic strength. For comparative purposes, solubility data as a function of pressure were colle cted on lysozyme. A linear dependence of lysozyme solubility with pressure was observed, with a 40% increase in apparent solubility after incubation f or seven days at 210 MPa and 25 degrees C. (C) 1999 Elsevier Science B.V. A ll rights reserved.