耐热木聚糖酶基因的克隆表达及其在酶解玉米芯木聚糖中的应用
发布时间:2018-10-25 08:29
【摘要】:为寻求具有耐热性能的木聚糖酶,笔者以嗜热网球菌(Dictyoglomus thermophilum)DSM3960的基因组为模板,克隆得到木聚糖酶基因xyn B-DT,该基因全长1 083 bp,共编码361个氨基酸,蛋白的理论分子量约为40ku。通过NCBI数据库比对发现该基因编码的蛋白质属于糖苷水解酶G11家族。实现大肠杆菌异源表达重组木聚糖酶Xyn B-DT,通过IPTG诱导,酶活达到30.6 U/mL。该重组木聚糖酶的最适温度为85℃,在60~80℃范围内均有较好温度稳定性,在60℃条件下保温2 h,酶活维持在90%以上,在90℃下保温2 h,酶活尚残余约50%;最适pH为6.5,在pH为5.0~7.5范围内保温24 h仍可保留约90%剩余酶活力。该酶以Beechwood木聚糖为底物,米氏常数(K_m)和最大反应速率(V_(max))值分别为5.63 mg/mL和1.572 mmol/(L·min~(-1))。以玉米芯木聚糖为底物,研究XynB-DT水解玉米芯木聚糖的条件及产物,结果显示在温度70℃、pH 6.0条件下酶解12 h,加酶量为400 U/g,最终酶解得率为44.3%,玉米芯木聚糖的水解产物主要以木二糖和木三糖为主,表明该木聚糖酶在低聚木糖制备方面具有较大应用潜能。
[Abstract]:In order to search for xylanase with heat resistance, the Xylanase gene xyn B-DT1 was cloned from the genome of (Dictyoglomus thermophilum) DSM3960 of Neisseria thermophilus. The total length of Xylanase gene 1 083 bp, encodes 361 amino acids, and the theoretical molecular weight of the protein is about 40ku. The protein encoded by this gene was found to belong to the glycoside hydrolase G11 family by NCBI database comparison. The recombinant xylanase Xyn B-DTwas expressed in Escherichia coli. The activity of the recombinant xylanase was 30.6 U / mL induced by IPTG. The optimum temperature of the recombinant xylanase is 85 鈩,
本文编号:2293165
[Abstract]:In order to search for xylanase with heat resistance, the Xylanase gene xyn B-DT1 was cloned from the genome of (Dictyoglomus thermophilum) DSM3960 of Neisseria thermophilus. The total length of Xylanase gene 1 083 bp, encodes 361 amino acids, and the theoretical molecular weight of the protein is about 40ku. The protein encoded by this gene was found to belong to the glycoside hydrolase G11 family by NCBI database comparison. The recombinant xylanase Xyn B-DTwas expressed in Escherichia coli. The activity of the recombinant xylanase was 30.6 U / mL induced by IPTG. The optimum temperature of the recombinant xylanase is 85 鈩,
本文编号:2293165
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