Solvent Deuterium Isotope Effect on the Binding of β-d-Galactopyranosyl Derivatives to β-Galactosidase (Escherichia coli, lac Z)

A value of 1.8 has been determined for (KI)HOH/(KI)DOD, the ratio of the values of KI for competitive inhibition of β-galactosidase by isopropyl β-d-thiogalactopyranoside in H2O and D2O. This is similar to the value of 1.7 for (Km)HOH/(Km)DOD, the ratio of the Michaelis constants determined for the β-galactosidase-catalyzed hydrolysis of 4-nitrophenyl β-d-galactopyranoside (Gal-OPNP) in H2O and D2O. The similarity of these solvent deuterium isotope effects suggests that the observed isotope effect on Km corresponds, mainly, to the isotope effect on the dissociation constant Kd for Gal-OPNP. The implications of these results for the interpretation of the solvent deuterium isotope effects on kcat and kcat/Km for β-galactosidase-catalyzed hydrolysis of Gal-OPNP is discussed.