【摘要】：為尋求具有耐熱性能的木聚糖酶,筆者以嗜熱網(wǎng)球菌(Dictyoglomus thermophilum)DSM3960的基因組為模板,克隆得到木聚糖酶基因xyn B-DT,該基因全長1 083 bp,共編碼361個氨基酸,蛋白的理論分子量約為40ku。通過NCBI數(shù)據(jù)庫比對發(fā)現(xiàn)該基因編碼的蛋白質(zhì)屬于糖苷水解酶G11家族。實現(xiàn)大腸桿菌異源表達(dá)重組木聚糖酶Xyn B-DT,通過IPTG誘導(dǎo),酶活達(dá)到30.6 U/mL。該重組木聚糖酶的最適溫度為85℃,在60~80℃范圍內(nèi)均有較好溫度穩(wěn)定性,在60℃條件下保溫2 h,酶活維持在90%以上,在90℃下保溫2 h,酶活尚殘余約50%;最適pH為6.5,在pH為5.0~7.5范圍內(nèi)保溫24 h仍可保留約90%剩余酶活力。該酶以Beechwood木聚糖為底物,米氏常數(shù)(K_m)和最大反應(yīng)速率(V_(max))值分別為5.63 mg/mL和1.572 mmol/(L·min~(-1))。以玉米芯木聚糖為底物,研究XynB-DT水解玉米芯木聚糖的條件及產(chǎn)物,結(jié)果顯示在溫度70℃、pH 6.0條件下酶解12 h,加酶量為400 U/g,最終酶解得率為44.3%,玉米芯木聚糖的水解產(chǎn)物主要以木二糖和木三糖為主,表明該木聚糖酶在低聚木糖制備方面具有較大應(yīng)用潛能。
[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，

本文編號：2293165