THE SOLID-STATE ENZYME-REACTIONS .4. HYDR OLYSIS OF BETA-INDOLEACRYLOYL-ALPHA-CHYMOTRYPSIN

Beta-Indoleacryloyl-alpha-chymotrypsin stable in aqueous solution at p H 5.0 is shown to be stable also in lyophylized state at low hydration for to 120 h, if the hydration degree does not exceed critical value (H(crit)) 135 mole H2O per mole of protein typical for other chymotryp sin-catalyzed solid-state and solvent-free reactions. However upon hyd ration above H(crit) the hydrolysis of acylenzyme does occur at t = t0 , the reaction to be considered as an enzyme-like process. In contrast to solid-state chymotrypsin reactions with SPPNA and PMSF very slow h ydrolysis of beta-indoleacryloyl-alpha-chymotrypsin took place at t > > t0 even at hydrations below the critical value. This reaction is pro bably a non-enzymatic general-base catalyzed hydrolysis. In this case the catalytic site of beta-indoleacryloyl-alpha-chymotrypsin can bind two water molecules of which one is a nucleophile and the other may be involved in the process of proton transfer from nucleophile to the le aving group, i.e. Ser 195 beta-oxygen atom of the catalytic site.