地衣芽胞杆菌碱性蛋白酶嗜碱突变体的特征分析  

Biochemical characterization of alkalophilic mutant of Bacillus licheniformis protease

作　　者：黄磊[1] 董自星[2] 金鹏[2] 王正祥[2] 路福平[1] 

机构地区：[1]天津科技大学生物工程学院工业发酵微生物教育部重点实验室,天津300457 [2]天津科技大学化工与材料学院生物化工系,天津300457

出　　处：《食品与发酵工业》2017年第6期34-40,共7页Food and Fermentation Industries

基　　金：国家自然科学基金面上项目(31370076);福建省自然科学基金(2016J01157)

摘　　要：地衣芽胞杆菌碱性蛋白酶是工业上重要的蛋白酶类,进一步提高其在碱性条件下的活力可改善其在洗涤剂工业方面的应用价值。该研究通过筛选获得一种在碱性条件下酶活水平显著提高的地衣芽胞杆菌B186来源的蛋白酶,并在枯草芽胞杆菌WB600中对其编码基因进行了成功克隆与表达,获得了重组菌WB600(pHYE209)。然后通过离子交换色谱和凝胶层析法,从重组菌的发酵液中纯化获得了重组碱性蛋白酶AprE209。对酶学性质的研究表明,重组酶Apr E209的最适作用pH为11.0,最适作用温度为50℃,在30~37℃下和pH12.0时仍具有很高的活力。进一步通过生物信息学的手段对其耐碱机制进行了初步解析。该嗜碱突变体具有进一步用于洗涤剂的潜在研究价值。Alkaline protease from Bacillus licheniformis is an important protease in the industry. Improving its ac- tivity and stability under alkaline pH conditions can broaden the range of applications in detergent industry. In this study, a high-alkaline protease from B. licheniformis B186 was screened, and its encoding gene was successfully cloned and expressed in B. subtilis WB600 to generate the recombinant bacterium WB600 （pHY-E209）. Recombi- nant alkaline protease AprE209 was then purified to apparent homogeneity from the supernatant of the recombinant bacterium using ion exchange chromatography and filtration chromatography. The optimum pH and temperature of re- combinant enzyme AprE209 were 11.0 and 50 ℃ , respectively. This recombinant enzyme also retained high activities at temperatures of 30 -37 ℃or pH 12.0. Furthermore, the mechanism of alkali-tolerance of AprE209 was analyzed by bioinformatics tools. The alkalophilic mutant of B. licheniformis protease obtained in this study had potential appli- cations in detergent industry.

关 键 词：蛋白酶 嗜碱性突变体 洗涤剂行业 酶学特征 

分 类 号：Q936[生物学—微生物学] TQ649[化学工程—精细化工]