摘要
运用基因工程技术,将克隆到的淀粉液化芽孢杆菌Bacillus amyloliquefaciens DSM 1061新型中性植酸酶基因(GenBank登录号为HM747163)构建到表达载体pET22b(+)上,并在E.coli BL21(DE3)中进行高效表达。电泳结果表明该酶分子质量约为42kD,目的蛋白占大肠杆菌可溶性蛋白30%左右。利用Ni-NTA琼脂糖凝胶进行亲和纯化,酶学性质研究表明,其最适温度为60℃,最适pH值为7.0,最大酶比活力为15U/mg。60℃时以植酸钠为底物的Km值为0.30mmol/L。
A new neutral phytase gene（phyc） from Bacillus amyloliquefaciens DSM 1061（GenBank： HM747163） was cloned into expression vector pET22b（＋）.The phytase was expressed in Escherichia coli BL21（DE3）.The molecule weight of the phytase protein was about 42 kD determined by SDS-PAGE.The expressed phytase was about 30% of the total soluble protein of E.coli.The recombinant protein was purified by 6×His-Tagged Protein Purification Kit.Optimal pH value and temperature of the phytase were 7.0 and 60 ℃,respectively.Under the optimum conditions,the activity was 15 U/mg.The Km value of the phytase for hydrolysis of sodium phytate under 60 ℃ was 0.30 mmol/L.
出处
《食品科学》
EI
CAS
CSCD
北大核心
2012年第21期153-156,共4页
Food Science
基金
国家自然科学青年科学基金项目(31101912)
江苏省自然科学基金面上项目(BK2011420)
江苏省普通高校自然科学研究项目(09KJB530011)
关键词
淀粉液化芽孢杆菌
高效表达
中性植酸酶
酶学性质
Bacillus amyloliquefaciens
overexpression
neutral phytase
enzymatic properties
