Carbonic anhydrase from Cyprinus carpio

Abstract

Carbonic anhydrase (CA) is the basic enzyme in the breathing function and catalyses the reversible hydration of CO2 to HCO3- and H+.A method is introduced to purify the carbonic anhydrase from erythrocytes of the carp Cyprinus carpio using an affinity gel (Sepharose 4B-L-tyrosine-p-aminobenzensulfonamide). This procedure resulted in a 314-fold purification of the enzyme. The purity of the enzyme was controlled with SDS-polyacrylamide gel electrophoresis, and the enzyme was observed as a single band. The CA has an esterase activity as well as a hydratase activity. The K-m and V-max values of the CA were calculated as 1.49 mM and 8.77 U/ml x min, respectively, using p-nitrophenylacetate as a substrate. The 150 values of inhibitors for the CA were determined by means of activity percentage diagrams. The values for I-50 were observed as 1.86 x 10(-5) M, 2.68 x 10(-5) M, 1.24 x 10(-5) M, 0.65 x 10(-5) M and 0.259 x 10(-5) M for sodium azide, sodium perchlorate, benzensulfonamide, dichlorbenzensulfonamide and acetazolamide, respectively.