Purification and characterization of carbonic anhydrase from liver of siraz (Capoeta umbla ), effects of some metals and pesticidies' on the enzyme activity

In this thesis , invitroeffects of some ions and pesticides on carbonicanhydrase (CA, carbonatehydrolyase, E.C. 4.2.1.1) enzym from Siraz (Capoeta umbla)liver tissue were investigated. Carbonicanhydrase enzyme was firstly purified from Siraz (Capoeta umbla) using by affinity gel chromatography (Sepharose-4B-L tyrosine-sulfanilamide) . As a result of affinity chromatography with a spescific activity of 721,0 EU / mg protein and 27.95 % efficiency, 120,709 fold purified. Sodiumdodecylsulfate-polyacrylamide gel (SDS-PAGE) electrophoresis was performed to check the purity of the purified enzyme and a single band was obtained. Sirase (C.umbla) livertissue in the kinetic studies for CA enzyme; Km and Vmax were calculated from Lineweaver-Burk plotsusing p-nitrophenylacetate substrate. Optimum pH in the K-Phosphatewas 7.0, optimum pH in the TRİS- SO4 was 8.5 , optimum ionicstrengths was 0.080 M Tris-SO4buffer, optimum temperature was 35° C, for the enzyme and stable pH was found in the K-phosphate buffer at pH 8.5. Theeffects of some metal ions and some pesticides on purified CA enzyme were investigated and Cu2+, Mn3+, Ba2+, Se2- ions which showeffect of inhibition and valves of of IC50 and Ki constantsare calculatedfor aldrın, dıeldrın, endosülfan, hexachlorbenzene, alpha- HCH pesticides.