重组海参溶菌酶工程菌发酵及表达产物纯化和性质的研究  

Study on Fermentation,Purification and Characterization of Recombinant Stichopus japonicus Lysozyme

作　　者：王丹[1] 丛丽娜[1] 谢三群[1] 张欢[1] 

机构地区：[1]大连工业大学生物工程学院,大连116034

出　　处：《生物技术通报》2011年第6期187-192,共6页Biotechnology Bulletin

基　　金：国家自然科学基金项目(31072224);辽宁省教育厅创新团队项目(LT2010012);大连市科技计划项目(2009J31SC034)

摘　　要：旨在研究重组海参i型溶菌酶(Stichopus japonicus lysozyme,SjLys)可溶性表达的发酵条件、纯化及生物学性质。采用摇瓶发酵的培养方式,分别从培养基组分和发酵条件两个方面对获得可溶性重组SjLys进行研究。表达产物通过亲和层析纯化以及凝胶过滤层析脱盐,获得了电泳纯为95%的重组SjLys蛋白。进一步对其生物学活性分析,结果表明该酶最适温度为45℃,最适pH值为6.8,对革兰氏阴性菌和革兰氏阳性菌均有广谱抑菌作用。上述结果将为海参溶菌酶进行产业化奠定应用基础。The project study mainly focused on fermentation of the genetically engineered bacteria to express the Stichopus japoni- cus lysozyme（ SjLys）, such as E. coil BL21 （DE3）pLysS/pET 32a （ ＋ ）SjLys, as well as purification and characterization of its product. The recombinant SjLys was first expressed successfully in solube form via optimization of fermentation conditions. The product was purified by Ni2＋ metal affinity chromatography（MAC） and then desahed by sephadex G-25 medium. The optimal pH and temperature acting on the substrate Micrococcus lysodeikticus was 6. 8 and 45℃ respectively. The antibacterial activity of the purified recombinant SjLys was analyed. It showed a broad antimicrobial spectrum against the tested strains including Gram-positive and Gram-negative bacteria. The study of SjLys could provide a basis for production of the sea cucumber lysozyme in a large scale.

关 键 词：可溶性 蛋白纯化 酶活性 溶菌酶 重组表达 

分 类 号：Q814[生物学—生物工程]