Agri Ibrahim Cecen University, Vocational Training School, Agri, Turkey.
Bioorg Med Chem. 2013 Mar 15;21(6):1522-5. doi: 10.1016/j.bmc.2012.08.018. Epub 2012 Aug 24.
Carbonic anhydrase (CA, EC: 4.2.1.1) was purified from sheep kidney by affinity chromatography on a Sepharose 4B-tyrosine-sulfanilamide column. By means of two consecutive procedures, the enzyme (sCA) was purified 227.61-fold with a yield of 60.75%, and a specific activity of 838.89U/mg proteins. The optimum temperature, ionic strength and pH were determined to be 35°C, 20mM and 8.5, respectively. The molecular weight determined by SDS-PAGE was found to be 29kDa. The kinetic parameters, KM and Vmax values were determined for the 4-nitrophenyl acetate (p-NpA) hydrolysis reaction. Some sulfonamides were tested as inhibitors against the purified CAs enzyme. The Ki constants for benzenesulfonamide (1), sulfanilamide (2), mafenide (3), 4-(2-aminoethyl) benzenesulfonamide (4), 4-methyl-benzenesulfonamide (5), 2-bromo-benzenesulfonamide (6), naphthalene-2-sulfonamide (7), 4-amino-6-chlorobenzene-1,3-disulfonamide (8) and saccharin (9) were in the range 1.348-69.31μM.
碳酸酐酶（CA，EC：4.2.1.1）通过亲和层析在 Sepharose 4B-酪氨酸-磺胺柱上从绵羊肾脏中纯化。通过两个连续的程序，该酶（sCA）被纯化了 227.61 倍，收率为 60.75%，比活为 838.89U/mg 蛋白。确定最佳温度、离子强度和 pH 值分别为 35°C、20mM 和 8.5。通过 SDS-PAGE 确定的分子量为 29kDa。确定了 4-硝基苯乙酸酯（p-NpA）水解反应的动力学参数 KM 和 Vmax 值。测试了一些磺胺类化合物作为对纯化的 CA 酶的抑制剂。苯磺酰胺（1）、磺胺（2）、甲灭酸（3）、4-（2-氨基乙基）苯磺酰胺（4）、4-甲基苯磺酰胺（5）、2-溴苯磺酰胺（6）、萘-2-磺酰胺（7）、4-氨基-6-氯-1,3-苯二磺酰胺（8）和糖精（9）的 Ki 常数在 1.348-69.31μM 范围内。
