| 15895688 |
Demir Y, Nadaroglu H, Demir N: Purification and characterization of carbonic anhydrase from bovine stomach and effects of some known inhibitors on enzyme activity. J Enzyme Inhib Med Chem. 2005 Feb;20(1):75-80.
Carbonic anhydrase (CA) was purified from four different cell localisation (outer peripheral, cytosolic, inner peripheral and integral) in bovine stomach using affinity chromatography with Sepharose-4B-L-tyrosine sulphanilamide. During the purification steps, the activity of the enzyme was measured using p-nitrophenyl acetate at pH 7.4. Optimum pH and optimum temperature values for all CA samples were determined, and their K (m) and V (max) values for the same substrate by Lineweaver-Burk graphics. The extent of purification for all CA localizations was controlled by SDS-PAGE. The K (m) values at optimum pH and 20 degrees C were 0.625 mM, 0.541 mM, 0.785 mM and 0.862 mM with p-nitro phenyl acetate, for all CA localizations. The respective V (max) values at optimum pH and 20 degrees C were 0.875 micromol/L min, 0.186 micromol/L min, 0.214 micromol/L min and 0.253 micromol/L min with the same substrate. The K (i) and I50 values for the inhibitors sulphanilamide, KSCN, NaN3 and acetazolamide were determined for all the CA localizations. |
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