C-13 NMR-STUDY OF CO2-HCO3-EXCHANGE CATALYZED BY HUMAN CARBONIC ANHYDRASE-C AT CHEMICAL-EQUILIBRIUM

The effects of human carbonic anhydrase C on the 13C nuclear magnetic resonance spectra of equilibrium mixtures of 13CO2 and NaH13CO3 were measured at 67.89 MHz. Enzyme‐catalyzed CO2‐HCO3− exchange rates were estimated from the linewidths of the resonances. The results show that: (a) the maximal exchange rates are larger than the maximal turnover rates; (b) the exchange is equally rapid with 1H2O or with 2H2O as solvents; (c) the exchange is equally rapid in the presence or in the absence of added buffers; (d) the apparent substrate binding is weaker than predicted if steady‐state Km values are assumed to represent substrate dissociation constants. The main conclusion concerning the catalytic mechanism of the enzyme is that the proton‐transfer processes which limit turnover rates in the steady state are not directly involved in CO2‐HCO3− exchange. In addition, the results suggest that CO2‐HCO3− interconversion takes place by a nucleophilic mechanism, such as a reversible reaction of zinc‐coordinated OH− with CO2. Copyright © 1979, Wiley Blackwell. All rights reserved