Kinetic behavior and some properties of carbonic anhydrase enzyme purified from gills of rainbow trout (Oncorhynchus mykiss) were studied at 4C. The purification steps included high-speed centrifugation, sepharose-4B-L tyrosine-sulfanilamide affinity gel chromatography and dialysis. Yield and specific activity of the enzyme were 40% and 55.56 EU / mg protein, respectively. The overall purification was 104.8-fold. The molecular mass was approximately estimated as 28 kDa by SDS polyacrylamide gel electrophoresis and as 27 kDa by gel filtration column chromatography. Optimal pH, stable pH and optimal temperature of the enzyme were 9.5, 8.2 in 0.025 M boric acid buffer and 17.5C, respectively. K-M and V-max values of the enzyme for the substrate (p-nitrophenylacetate) were 8.13 mM and 2.10 mu mol / mg protein / min, respectively. The dissociation constant of the enzyme inhibitor complex (K-i) value was 3.93 +/- 0.65 mu M for sulfanilamide and sulfanilamide inhibited the enzyme in a noncompetitive manner. Data from present study showed that optimum kinetic properties of gill CA enzyme were different from other vertebrate CA's.