Human carbonic anhydrase IX (CA IX) is an integral membrane protein and a member of the a class of carbonic anhydrases that includes the human and animal enzymes. We have prepared a truncated, recombinant form of human CA IX of 255 residues consistent with full-length hv man CA II, among the most efficient of the carbonic anhydrases. Catalysis by and inhibition of this form of human CA IX has been investigated using stopped-flow spectrophotometry and O-18 exchange measured by mass spectrometry. In kinetic constants for the hydration of CO,, CA IX closely resembled CA II with maximal proton transfer-dependent O-18 exchange near I mus(-1) and k(cat)/K-m near 55 muM(-1) s(-1). Human CA IX was very strongly inhibited by three classic sulfonamides and cyanate, with inhibition constants that are close to those for CA Il.