大豆磷酸烯醇式丙酮酸磷酸酯酶研究 Ⅲ、非专一性酸性磷酸酯酶的纯化与特性

Study on soybean phosphoenolpyruvate phosphatase(PEPP)Ⅲ.Purification and characterization of nonspecific acid phosphatase

从大豆叶片中分离能水解磷酸烯醇式丙酮酸(PEP)的酸性磷酸酯酶,通过硫酸胺分部(20%～50%饱和度)沉淀、DEAE纤维素层析、刀豆球蛋白琼脂糖凝胶亲和层析将酶纯化了422.47倍,活性达78.16U/mg蛋白。该酶对PEP专一性不强,Km为1.09mmol/L(PEP),最适pH5.8,在pH4.8～7.0范围内及60℃以下较稳定,水解PEP的活性被Mg2+、Mn2+激活,F、Cu2+、Zn2+、PO43、MoO42及3磷酸甘油酸(3PGA)、三磷酸腺苷ATP等代谢物抑制,受异柠檬酸等有机酸影响较小。

An acid phosphatase hydrolyzing phosphoenolpyruvate（PEP）has been isolated from soybean leaves and purified 422.47-fold,to a final specific activity of 78.16 U/mg proteins, through Ammonium sulfate fraction precipitation（20% -50 % saturation）,DEAE-cellulose chromatography and Concanavalin A sepharose affinity chromatography. This enzyme shows low specificity for phosphoenolpyruvate（PEP）; its Km is 1.09 mmol/L and optimum pH15.8. It is relatively stable at the pH ranging from 4.8 to 7.0 and at temperature below 60 ℃. The enzyme activity is activated by Mg^2＋ ,Mn^2＋ and inhibited by F^- ,Cu^2＋ ,Zn^2＋ ,pO4^3-,MoO4^2- and some metabolites such as 3-PGA,other than organic acids such as isocitrate.