H 2 CO 3  as substrate for carbonic anhydrase in the dehydration of HCO 3 .
Carbonic anhydrase, a metalloenzyme containing one zinc atom per protein molecule of molecular weight 30,000, catalyzes the interconversion of CO(2) and HCO(3) (-) in solution. The rate of catalysis, among the fastest known, is pH-dependent, with a pK(Enz) near neutral. Arguments are presented to show that: (i) only the high-pH form of the enzyme is active both for the hydration and dehydration reactions (ii) at high pH there is an H(2)O ligand on the metal (not an OH(-) as is often argued), and (iii) the substrate for the dehydration reaction is the neutral H(2)CO(3) molecule. The arguments are based on data in the literature on the nuclear relaxation rates of Cl(-) ions and water protons in solutions of carbonic anhydrase, on strict application of the principle of microscopic reversibility, and on kinetic considerations. It has been argued that H(2)CO(3) cannot be the substrate for the dehydration reaction because the observed CO(2) production rate is somewhat faster than the maximum rate at which H(2)CO(3) molecules can diffuse to the active site of the enzyme. However, current models that consider HCO(3) (-) as the substrate implicity require that protons diffuse to the enzyme at an even greater rate, well outside the limitations imposed by diffusion. We consider two mechanisms to obviate the diffusion limitation problem, and conjecture that at high substrate concentration, H(2)CO(3) reaches the active site by collision with the enzyme molecule, and subsequent surface diffusion to the active site. At lower substrate concentrations, corresponding to [HCO(3) (-)] <1 mM, generation of H(2)CO(3) molecules near the enzyme by the recombination reaction H(+) + HCO(3) (-) --> H(2)CO(3) can supply an adequate flux of substrate to the active site.
D001639:Bicarbonates; D001665:Binding Sites; D002245:Carbon Dioxide; D002256:Carbonic Anhydrases; D002384:Catalysis; D002712:Chlorides; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008433:Mathematics; D008956:Models, Chemical; D011522:Protons
D001639:Bicarbonates; D002712:Chlorides; D011522:Protons; D002245:Carbon Dioxide; D002256:Carbonic Anhydrases