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OALib Journal期刊
ISSN: 2333-9721
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来源于地衣芽胞杆菌的角蛋白酶在大肠杆菌中的表达、理化性质及其发酵优化
DOI: 10.3724/SP.J.1145.2013.00997, PP. 997-1002
Keywords: 角蛋白酶,地衣芽胞杆菌,大肠杆菌,重组表达,酶学性质,发酵优化
Abstract:
为了提高角蛋白酶在大肠杆菌中的胞外表达量并了解其酶学性质，从一株具有羽毛降解能力的地衣芽胞杆菌（bacilluslicheniformisbbe11-1）中克隆获得全长为1140bp的角蛋白酶基因ker，将去掉自身信号肽全长为1053bp的ker基因与质粒pet-22b(+)进行连接并转化到大肠杆菌bl21宿主中获得重组菌株.此后，采用疏水苯基层析柱对所表达的角蛋白酶进行纯化，获得分子量（mr）大小约32×103的重组角蛋白酶.酶学性质研究表明，该重组角蛋白酶最适温度为50℃，最适ph值为10，属于丝氨酸蛋白酶，金属离子mg2+和k+对酶活力有促进作用，而mn2+对其有抑制作用.经优化后，该酶胞外酶活在诱导温度20℃、iptg0.05mmol/l、甘氨酸浓度150mmol/l以及酵母粉浓度60g/l的条件下可达460.2u/ml.研究表明，优化培养条件可以较好地提高角蛋白酶在大肠杆菌中的表达量，结果为利用基因工程菌生产角蛋白酶奠定了基础.
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