STUDY OF THE SOLID-STATE ENZYME-REACTIONS .3. IRREVERSIBLE INACTIVATION OF ALPHA-CHYMOTRYPSIN BY BENZYLSULFONYL FLUORIDE

In solvent-free solid mixture of alpha-chymotrypsin and benzylsulfonyl fluoride (PMSF) the irreversible inactivation of the enzyme's activity takes place when hydration (H) of the protein molecule exceeds a certain critical value H(crit) = 127 moles of H2O per mole of alpha-chymotrypsin. This value is approximately equal to the H(crit) obtained for the solid-state chymotrypsin-catalysed hydrolysis of a substrate, N-succinyl-L-phenylalanine p-nitroanilide. The shape of the inactivation isotherm confirms the phase transition at H(crit) and demonstrates that the main mass of the protein (up to 75%) is activable, but heterogeneous with respect to the hydration level necessary for switching on the catalytic activity of alpha-chymotrypsin.