土曲霉金色变种AT8951菊粉酶的纯化和性质的研究

PURIFICATION AND PROPERTIES OF INULINASE FROM ASPERGILLUS TERREUS VAR. AUREUS AT8951

土典霉金色变种AT8951菊粉酶粗酶液经硫酸铵分段沉淀、DEAE Cellulose DE32离子交换、超滤、Sephadex G-150凝胶过滤和FPLC,获得两个菊粉酶组分EⅠ和EⅡ,经分析型FPLG和PAGE鉴定为单一纯和分析纯。EⅠ分子量为66KD,最适作用温度和pH分别55℃和5.8;EⅡ分子量为56KD,最适作用温度为57℃,最适pH为6.0。EⅠ和EⅡ皆为糖蛋白,多糖含量分别为24.7％和22％,都属于内切酶。本文还对EⅠ和EⅡ的Km值和I/s值,温度、pH、离子对酶活作用的影响等进行了研究。

The inulinase from Aspergillus terreus var. aureus crude enzyme preparation was purified to two components, designated as EI and EII, by means of (NH4)2SO4 fractionation, DEAE Cellulose DE32 ion exchange chromatography, ultrafiltration, Sephadex G-150 gel filtration and FPLC. EI and EII were demonstrated to be homogeneous by analytical FPLC and PAGE. El had a molecular weight of 66KD, an optimal temperature of 55℃ and an optimal pH of 5.8. EII had a molecular weight of 56KD, an optimal temperature of 57℃ and an optimal pH of 6.0. Both EI and EII were glycoproteins with saccharide contents of 24.7% and 22% respectively and belonged to the endo-inulinase. The Km values and I/S ratios of EI and EII and the effects of temperature, pH, various metal ions on inulinase activity were also studied.