P. W. Taylor, RW King, and A. S. V. Burgen abstract: The kinetics of complex formation between aromatic sulfonamides and human carbonic anhydrases B and C have been examined over the pH range, 5.0-10.8 by stopped-flow kinetic measurements of fluorescence quenching. The apparent bimolecular association rate constant is highly pH dependent whereas the dissociation rate shows no variance with pH. The kinetic constants and their pH dependence are similar for the Zn and Co (II) forms of theenzyme. Correlation between the visible dd absorption spectra of the Co (II) enzyme and the association kinetics suggest that the pH dependence of the association process is governed by a proton-dependent equilibrium between different coordination forms of the enzyme. Combination appears to occur between the neutral sulfonamide species and the coordination form of carbonic anhydrase which predominates at alkaline pH. The pH-insensitive dissociation

/Aromatic and heterocyclic sulfonamides form a group of specific inhibitors of the metalloenzyme, carbonic anhy-drase, possessing a wide range of affinity constants (Maren, 1967; Taylor et al., 1970). The crystal structure of a human carbonic anhydrase C-acetoxymercurisulfanilamide complex shows that the sulfonamido group resides in close vicinity to the coordination sphere of the metal (Fridborg et al., 1967). Replacement of the single Zn atom in the enzyme by Co generate a visible dd spectrum characteristic of the coordinated ligands and their geometric arrangement around the metal (Lindskog, 1963; Lindskog and Nyman, 1964; Coleman, 1968). Changes in the Co (II) spectrum accom-panying sulfonamide binding further support the concept that the sulfonamide may function as a metalloligand. Fluorescence enhancement observed with the sulfonamide, dansylamide, suggests that theessential aromatic ring of this inhibitor is bound in a relatively hydrophobic environ-ment in the protein (Chenand Kernohan, 1967). Thus stabilization of the complex may be conferred through