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Hot Biological Catalysis: Isothermal Titration Calorimetry to Characterize Enzymatic Reactions
Published on: April 4, 2014
The catalytic properties of human carbonic anhydrase IX
1Department of Pharmacology and Therapeutics, University of Florida College of Medicine, Gainesville, Florida 32610-0267, USA.
Abstract:
Human carbonic anhydrase IX (CA IX) is an integral membrane protein and a member of the alpha 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 human 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 18O exchange measured by mass spectrometry. In kinetic constants for the hydration of CO2, CA IX closely resembled CA II with maximal proton transfer-dependent 18O exchange near 1 micros(-1) and kcat/Km near 55 microM(-1) x 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 II.
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