Manganaro F, Gaudette Y, Pombo-Gentile A, Singh K, Rakhit S
Bio-Méga, Inc., Laval, Québec, Canada.
Prostaglandins. 1988 Dec;36(6):859-74. doi: 10.1016/0090-6980(88)90062-7.
Leukotriene A4 epoxide hydrolase from dog lung, a soluble enzyme catalyzing the hydrolysis of leukotriene A4 (LTA4) to leukotriene B4 (LTB4) was partially purified by anion exchange HPLC. The enzymatic reaction obeys Michaelis- Menten kinetics. The apparent Km ranged between 15 and 25 microM and the enzyme exhibited an optimum activity at pH 7.8. An improved assay for the epoxide hydrolase has been developed using bovine serum albumin and EDTA to increase the conversion of LTA4 to LTB4. This method was used to produce 700 mg of LTB4 from LTA4 methyl ester. The partial by purified enzyme was found to be uncompetitively inhibited by divalent cations. Ca+2, Mn+2, Fe+2, Zn+2 and Cu+2 were found to have inhibitor constants (Ki) of 89 mM, 3.4 mM, 1.1 mM, 0.57 mM, and 28 microM respectively Eicosapentaenoic acid was shown to be a competitive inhibitor of this enzyme with a Ki of 200 microM. From these inhibition studies, it can be theorized that the epoxide hydrolase has at least one hydrophobic and one hydrophilic binding site.
犬肺白三烯A4环氧水解酶是一种可催化白三烯A4（LTA4）水解为白三烯B4（LTB4）的可溶性酶，通过阴离子交换高效液相色谱法对其进行了部分纯化。酶促反应遵循米氏动力学。表观米氏常数（Km）在15至25微摩尔之间，该酶在pH 7.8时表现出最佳活性。利用牛血清白蛋白和乙二胺四乙酸（EDTA）开发了一种改进的环氧水解酶检测方法，以提高LTA4向LTB4的转化率。该方法用于从LTA4甲酯生产700毫克LTB4。发现部分纯化的酶受到二价阳离子的非竞争性抑制。钙离子、锰离子、亚铁离子、锌离子和铜离子的抑制常数（Ki）分别为89毫摩尔、3.4毫摩尔、1.1毫摩尔、0.57毫摩尔和28微摩尔。二十碳五烯酸被证明是该酶的竞争性抑制剂，Ki为200微摩尔。从这些抑制研究可以推测，环氧水解酶至少有一个疏水结合位点和一个亲水结合位点。
