Molecular basis of the oxygen exchange from CO2 catalyzed by carbonic anhydrase III from bovine skeletal muscle.
The exchange of 18O from CO2 to H2O in aqueous solution is caused by the hydration-dehydration cycle and is catalyzed by the carbonic anhydrases. In our previous studies of 18O exchange at chemical equilibrium catalyzed by isozymes I and II of carbonic anhydrase, we observed simple first-order depletion of 18O from CO2 with the 18O distribution among the species C18O18O, C16O18O, and C16O16O described by the binomial expansion (i.e., a random distribution of 18O). Using membrane-inlet mass spectrometry, we have measured 18O exchange between CO2 and H2O catalyzed by native zinc-containing and cobalt(II)-substituted carbonic anhydrase III from bovine skeletal muscle near pH 7.5. The distributions of 18O in CO2 deviate from the binomial expansion and are accompanied by biphasic 18O-exchange patterns; moreover, we observed regions in which 18O loss from CO2 was faster than 18O loss from HCO3-. These data are interpreted in terms of a model that includes 18O loss from an enzyme-substrate or intermediate complex. We conclude that more than one 18O can be lost from CO2 per encounter with the active site of isozyme III, a process that requires scrambling of oxygens in a bicarbonate-enzyme complex and cycling between intermediate complexes. This suggests that the rate of dissociation of H2(18)O (or 18OH-) from isozyme III is comparable to or faster than substrate and product dissociation.
D000818:Animals; D002245:Carbon Dioxide / Q000378:metabolism*; D002256:Carbonic Anhydrases / Q000378:metabolism*; D002417:Cattle; D007553:Isotope Labeling / Q000379:methods; D007700:Kinetics; D013058:Mass Spectrometry; D009132:Muscles / Q000201:enzymology*; D010103:Oxygen Isotopes
D010103:Oxygen Isotopes; D002245:Carbon Dioxide; D002256:Carbonic Anhydrases