SOLVENT ISOTOPE EFFECTS ON A HYDROLYSIS REACTION CATALYZED BY SUBTILISIN AND ITS N155G MUTANT - FAILURE OF THE PROTON INVENTORY METHOD TO REPORT HYDROGEN-BONDING INTERACTIONS IN THE OXYANION HOLE

Proton inventories (solvent isotope effects in H2O-D2O mixtures) are not detectably different for the hydrolysis of an anilide substrate catalysed by subtilisin and its N155G mutant, despite the fact that this mutation removes a critical hydrogen-bonding interaction supplied by Asn-155 in the oxyanion hole of the enzyme, which stabilizes the transition state of this reaction by an amount equivalent to two orders of magnitude in reaction rate.