pH-Dependent Structure and Dynamics of the Catalytic Domains of Human Carbonic Anhydrase II and IX

Extensive computer simulation studies have been carried out to probe the pH-dependent structure and dynamics of the two most efficient isoenzymes II and IX of human carbonic anhydrase (HCA) that control the pH in the human body. The equilibrium structure and hydration of their catalytic domains are found to be largely unaffected by the variation of pH in the range studied, in close agreement with the known experimental results. In contrast, a significant effect of the change in pH is observed for the first time on the local electrostatic potential of the active site walls and the dynamics of active site water molecules. We also report for the first time the free energy and kinetics of coupled fluctuations of orientation and protonation states of the well-known His-mediated proton shuttle (His-64) in both isozymes at pH 7 and 8. The transitions between different tautomers of in or out conformations of His-64 side chain range between 10⁹ and 10⁶ s^-1 depending on pH. Possible implications of these results on conformation-dependent pK_a of His-64 side chain and its role in driving the catalysis toward hydration of CO₂ or dehydration of HCO₃^- with varying pH are discussed.