Carbon isotope effect on dehydration of bicarbonate ion catalyzed by carbonic anhydrase.
The carbon-13 kinetic isotope effect on the dehydration of HCO3- by bovine carbonic anhydrase has been measured. To accomplish this, bicarbonate was added to a buffer solution at pH 8 containing carbonic anhydrase under conditions where purging of the product CO2 from the solution is rapid. Measurement of the isotopic composition of the purged CO2 as a function of the concentration of carbonic anhydrase permits calculation of the isotope effect on the enzymic reaction. The isotope effect on the dehydration is k12/k13 = 1.0101 +/- 0.0004. This effect is most consistent with a ping-pong mechanism for carbonic anhydrase action, in which proton transfer to or from the enzyme occurs in a step separate from the dehydration step. Substrate and product dissociation steps are at least 2-3-fold faster than the hydration/dehydration step.
D000818:Animals; D001639:Bicarbonates / Q000378:metabolism*; D002247:Carbon Isotopes; D002256:Carbonic Anhydrases / Q000097:blood*; D002417:Cattle; D004912:Erythrocytes / Q000201:enzymology; D007553:Isotope Labeling / Q000379:methods; D007700:Kinetics; D008433:Mathematics
D001639:Bicarbonates; D002247:Carbon Isotopes; D002256:Carbonic Anhydrases