摘要
金属硫蛋白(MT)作为一类广泛存在并高度可诱导的多功能内源性蛋白,具有重金属解毒、抗辐射、清除自由基等功能,近年来已成为研究和开发的热点。为解决MT原料来源的瓶颈问题,本文以一株海洋源MT重组巴斯德毕赤酵母菌株(GS-115-MT)为研究对象,对其进行培育条件(温度、pH、溶氧、接种量)、诱导表达条件(溶氧、甲醇浓度、菌株生产状况、诱导时间、诱导金属及浓度)综合优化,以获得MT高效表达最佳工艺。结果显示,GS-115-MT的最佳生长条件为培育温度30℃,培养基初始pH为9,摇床转速240r·min^-1,接种量4%;最佳MT诱导发酵条件为摇床转速为220r·min^-1,甲醇浓度为5%,菌液OD600为9.7,诱导24h,Cu^2+诱导浓度50μmol·L^-1。在以上优化条件下,MT表达量达0.328mg·mL^-1。本结果可为海洋源MT的规模化工业生产提供一定理论依据。
Metallothionein(MT),as a kind of multifunctional endogenous protein,has been widely used in medicine, cosmetics,health food additives,environmental protection,etc.It has several functions,e.g.maintaining dynamic balance of metal content in organisms,detoxifying heavy metals,anti-radiation,and scavenging activity of free radicals.In this study,in order to solve the bottleneck problem of the sources of MT,Pichia pastoris recombinant strain(GS-115-MT) was selected as the research object,and its induction(temperature,pH,dissolved oxygen and inoculation amount) and expression conditions(dissolved oxygen,methanol concentration,OD600,induction time,induction metal ions and concentration)wrere optimized comprehensively to obtain the best way to express metallothionein efficiently.The results showed that the optimum conditions of GS-115-MT growth were as follows,incubation temperature 30℃,initial pH value 9, rotation speed 240r·min^-1 and inoculation volume 4%.The optimum conditions for MT induction were as follows,rotation speed 220r·min^-1,formaldehyde concentration 5%,bacterial liquid OD600 9.7,induction time 24h and induction metal 50μmol·L^-1 Cu^2+,when the MT expression was 0.328mg·mL^-1.The results of this study can provide a theoretical basis for large-scale industrial production of marine source MT.
作者
阎光宇
余蕾
何建林
孙继鹏
YAN Guangyu;YU Lei;HE Jianlin;SUN Jipeng(Xiamen Ocean Vocational College,Xiamen 361102,China;Collaborative Innovation Center for Marine Biotechnology,Higher Vocational College,Xiamen 361100,China;Third Institute of Oceanography,Ministry of Natural Resource,Xiamen 361005,China)
出处
《生物产业技术》
2019年第5期64-70,共7页
Biotechnology & Business
基金
厦门海洋职业技术学院自然科学育青项目(KYZY201807)
厦门海洋研究开发院共建项目(K190102)
福建省科技计划引导性项目(2017N0016)
关键词
金属硫蛋白
重组酵母菌株
发酵
诱导
metallothionein
recombinant Pichia pastoris
fermentation
induction
