ACTIVATION AND INHIBITION OF BOVINE CARBONIC ANHYDRASE-III BY DIANIONS

We have found that many dianionic species, at millimolar concentrations, significantly activate or inhibit the bovine carbonic anhydrase III-catalyzed hydration of CO2. Dianionic species such as HPO4(2-) and SO3(2-), with pK(b) values near 7, are activators, whereas weakly basic species such as SO4(2-) act as inhibitors. Both activation and inhibition are partial hyperbolic in nature and do not appear to compete with monoanionic linear inhibitors like N3-. Our kinetic data are consistent with a formal mechanism of action for carbonic anhydrase III that is directly analogous to that of carbonic anhydrase II, in which Lys-64 of carbonic anhydrase III can act as an intramolecular H+ transfer group during CO2 hydration. Our data suggest that dianionic inhibitors depress the rate of H+ transfer during turnover by stabilizing the protonated form of Lys-64. We postulate that dianionic activators enhance the rate of a rate-limiting H+ transfer step in the mechanism, probably by acting directly as H+ acceptors.