【摘要】 采用同源建模的方法构建拟青霉β-1,3(4)-葡聚糖酶的三维结构。通过对其活性位点及表面氨基酸残基侧链的分析,利用氨基载体Sepabeads EC-HA共价固定化葡聚糖酶,优化固定化条件,比较固定化酶与游离酶的酶学参数。结果表明:m(酶粉):m(载体)=1.2:1、温度40～45℃、固定化时间8h,固定化效果最好。蛋白结合率可达91.7%,酶活回收率达87.6%,固定化酶最适温度、热稳定性、pH值稳定性和批次使用稳定性均得到明显提高。更多还原

【Abstract】 The three-dimensional structure of β-1,3(4)-glucanase from Paecilomyces sp.FLH30 was constructed by means of homology modeling using the crystal structure of endo-β-1,3(4)-glucanase from Phanerochaete chrysosporium as a template,and its active site and side chains of surface amino acid residues were analyzed.Sepabeads EC-HA as a carrier of amino groups was used for the covalent immobilization of this enzyme and immobilization conditions were optimized.Meanwhile,enzymatic characteristics of free and immobilized β-1,3(4)-glucanase were compared.The best immobilization results were obtained under the conditions: enzyme/carrier mass ratio1.2:1,temperature 40—45 ℃,and immobilization time 8 h.Under these conditions,the protein binding rate was 91.7% and the activity recovery was 87.6%.The optimum temperature,thermal stability,pH stability and operational stability of immobilized glucanase were all improved when compared to free glucanase.更多还原