摘要
为了促进重组淀粉分支酶在枯草芽孢杆菌（Bacillus subtilis WB600）中的胞外表达，对重组淀粉分支酶在摇瓶水平上进行了发酵优化。首先，以胞外酶活力为指标，先后考察了初始培养基、pH、培养温度和培养时间4种因素对胞外表达的影响；然后，通过两阶段温度策略和助剂的添加进一步提高胞外表达水平。结果表明，当发酵培养基为TB、pH为7．5时，25℃培养24h后胞外酶活力达到19．9U／mL；同时，菌体生长最适温度和酶分泌的最适温度是不一致的，通过两阶段温度控制策略（即0—12h控制温度30℃，12h后温度调至25℃），胞外酶活力与单一温度条件下的最高水平（25℃）相比提高了1．4倍，达到了47．1U／mL；在此基础上，进一步添加0．05％的吐温-80，将胞外酶活进一步提高40％，达到65．8U／mL。
The fermentation conditions were optimized in the shake flask in order to further enhance the extracellular expression of recombinant Starch Branching Enzyme （SBE） by Bacillus subtilis WB600. Effects of four factors including culture media, initial pH, temperature and culture time were firstly investigated in succession on extracellular expression with extracellular enzyme activity as an index. The level of extracellular expression was further improved by a two-staged temperature strategy. The results showed that the extracellular SBE activity was 19.9 U/mL while the culture was performed in TB medium, pH7.5, at 25℃ for 24 h. Meanwhile the optimum temperature for cell growth is inconsistent with that for secretion. By the two-staged temperature strategy （temperature was controlled at 30 ℃ for the first 12 h and then switched to 25 ℃till the end of the fermentation） , the extracellular activity reached 47.1 U/ mL, increased by 140% compared to the constant temperature operation at 25 ℃. It was further increased by 40% to 65.8 U/mL after addition of Tween-80（0.05% , w/v）.
出处
《食品与发酵工业》
CAS
CSCD
北大核心
2016年第8期19-24,共6页
Food and Fermentation Industries
基金
高校博士点基金专项课题优先发展领域课题(20130093130001)
中国博士后科学基金-面上资助(2014M560394)
江苏省博士后科研资助计划(1401100C)
江苏省科技支撑计划-农业部分(BE2014305)
关键词
淀粉分支酶
胞外表达
两阶段温度控制策略
发酵优化
starch branching enzyme （SBE）
extracellular expression
two-staged temperature strategy
optimization of fermentation
