菜豆幼苗EPSP合成酶的分离纯化和它的部分性质

利用硫酸铵分级沉淀，SephedexG-50凝胶柱层析，FPLCMono-Q和磷酸纤维素离子层析法从菜豆幼苗中分离提纯了EPSP合成酶。该酶被纯化2961.6倍，比活性达到6219.4nmolmg-1蛋白min-1。该酶分子量经SDS-PAGE检测为51kD,等电点为pH5.7,酶促反应最适pH7.5,最适温度45℃。6.2μmol/L的除草剂草甘膦能抑制EPSP合成酶活性的50%。

Purification of EPSP Synthase from Bean Seedlings and Its Properties

EPSP synthase from bean seedlings was purified by sequential ammonium-sulphate precipitation, Sephadex G-50 (Fig.1) and FPLC Mono-Q chromatography (Fig.2) and substrate elution from cellulose phosphate column (Fig.3). The degree of purification was 2 961.6 fold. The specific activity was 6 219.4 nmol mg-1 protein min-1 (Table 1). The molecular weight was determined to be 51 kD by SDS-polyacrylamide gel eleetrophoresis (Fig. 4). The pH optimum (Fig.6), temperature optimum (Fig.7) and isoelectric point (Fig.5) of the purified EPSP synthase were 7.5, 45℃ and pH 5.5, respectively. Enzyme activity was inhibited approximately 50% by herbicide glyphosate 6.2 μmol/L (Fig.8).