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首先从菠菜叶片中纯化了乙醇酸氧化酶(GO)。通过鉴定反应中氧的消耗以及反应产物 H2O2的生成,证实菠菜 GO 具有氧化光呼吸途径中间代谢物甘油酸的活性。该氧化活性依赖于辅因子 FMN和 FAD,而不依赖核黄素和光黄素;其最适反应pH值为 8.0,Km(甘油酸)值为 7.14mmol/L,kcat值为1.04s-1,活化能为17.29kJ/mol;草酸和丙酮酸对该氧化活性有明显的抑制作用,其中前者为典型的竞争性抑制。进一步通过两底物竞争作图表明:菠菜叶片 GO 氧化甘油酸反应和氧化乙醇酸反应为同一活性中心所催化。菜叶片 GO 氧化甘油酸反应和氧化乙醇酸反应为同一活性中心所催化。
Glycolate oxidase (GO) was first purified from spinach leaves. Through the identification of oxygen consumption in the reaction and the formation of the reaction product H2O2, spinach GO was confirmed to have the activity of oxidizing the intermediate metabolite glycerate of the photorespiration pathway. The oxidative activity was dependent on the cofactors FMN and FAD, independent of riboflavin and leupeptin. The optimum reaction pH was 8.0, Km value was 7.14mmol / L, kcat value was 1.04s-1, The activation energy was 17.29kJ / mol; oxalic acid and pyruvate significantly inhibited the oxidation activity, of which the former was a typical competitive inhibition. Further mapping of the two substrates showed that GO oxidation and glycolate reaction of spinach leaves were catalyzed by the same active site. GO leaf oxidation and glycerate oxidation of glycolate reaction for the same active center catalysis.