沙田柚皮过氧化物酶的纯化及酶学性质的研究
Study on purification and properties of peroxidase from shatian pomelo
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摘要: 为了获得一种富含过氧化物酶(POD)的植物,本研究利用循环催化流动分析法(RCFA)初步筛选了15种植物样品,对样品中POD活性最高的沙田柚皮粗酶液进行了硫酸铵盐析和Sephadex G-200凝胶层析纯化,并对已纯化的POD进行了酶学性质研究。得到的结果表明:每克沙田柚皮中POD可达15.08U,纯化后的比活力可达313.29U/mg,纯化倍率为26.22倍,POD最终产率为43.20%;优化得到的POD最适温度和pH分别为55℃和5.5,在最适条件下测得该酶以H2O2为底物的米氏常数Km为0.099mmol/L,以愈创木酚(GA)为底物的米氏常数Km为0.864mmol/L。Abstract: In order to get a plant which was rich in peroxidase (POD ) , 15 species of plant were preliminary investigated by recirculating-catalysis flow analysis, and the Shatian pomelo peels peroxidase (SPP) whose enzymatic activity was the highest was purified by using ammonium sulfate grade precipitation and Sephadex G -200 gel chromatography. The result showed that:the POD activity was up to 15.08U in each gram of Shatianyou peels, the specific activity of purified SPP was up to 313.29U/mg, purification factor of 26.22 and yield of 43.20%. The optimum temperature and pH of SPP was 55℃ and 5.5 respectively. Its apparent Km for hydrogen peroxide and guaiacol was 0.099mmol/L and 0.864mmol/L under the optimal condition respectively.