重组水稻几丁质酶的发酵生产、纯化及酶学性质  

Production, Purification and Characteristics of the Recombinant Chitinase from Rice

作　　者：阎瑞香[1] 吴仲[1] 侯建华[2] 丁东风[2] 关文强[1] 李明刚[2] 

机构地区：[1]国家农产品保鲜工程技术研究中心,天津市农产品采后生理与贮藏保鲜重点实验室,天津300384 [2]南开大学分子生物学研究所,生物活性材料教育部重点实验室,天津300071

出　　处：《农业生物技术学报》2008年第1期148-153,共6页Journal of Agricultural Biotechnology

基　　金：天津市应用基础研究项目(No.06YFJMJC12200)资助

摘　　要：用7L发酵罐对毕赤酵母(Pichia pastoris)工程菌株NB01进行了重组几丁质酶的诱导表达,通过提高通气量和调整搅拌转速,使溶氧维持在30%以上,NB01可在发酵84h内达到产酶高峰,酶活最高可达48.0795U/mL。利用中空纤维超滤膜对发酵液浓缩、提纯,再经过Sephadex G-100柱层析后得到单一组分的重组几丁质酶,酶纯度提高了7.9961倍。重组几丁质酶在60℃以下具有良好的热稳定性,在pH2￣6范围内有较强的耐受力,在4℃条件下贮藏具有较好的稳定性。In a 7 L fermentor, production conditions of engineering strain Pichia pastoris NB01 containing chitinase gene were investigated. The maximum chitinase activity could reach 48.0795 U/mL at 84 h while dissolution oxygen （DO） level maintaining higher than 30% by enhancing aeration and changing agitation rate. The fast purification of recombinant chitinase was conducted with Hol- low fiber UF membrane and Sephadex G-100, and SDS-PAGE analysis indicated that there was a single band with the right molecular weight corresponding to the chitinase, so purification multiple of chitinase was enhanced 7.9961. Furthermore, the recombinant en- zyme was stable at 60 ℃ and pH 2-6. And it was rather stable for a long time when stored at 4℃.

关 键 词：重组几丁质酶 基因工程菌 发酵 纯化 

分 类 号：S182[农业科学—农业基础科学]