摘要
缺氧条件下嗜乙酰乙酸棒杆菌Corynebacterium acetoacidophilum ATCC13870生长停滞,却能够代谢葡萄糖产生以乳酸和琥珀酸为主的有机酸。采用以sacB基因为反向筛选标记的同源重组染色体基因敲除系统,敲除嗜乙酰乙酸棒杆菌的乳酸脱氢酶基因,得到的Δldh菌株CCTCC NO.M20122041在缺氧条件下不产乳酸,葡萄糖消耗速率降低了29.3%,产琥珀酸和乙酸浓度分别提高45.6%和182%;NADH/NAD+值小于1(约0.7);磷酸烯醇式丙酮酸羧化酶和乙酸激酶的比酶活分别提高84%和12倍。说明嗜乙酰乙酸棒杆菌中乳酸合成途径的阻断驱使了琥珀酸和乙酸代谢途径加强,推测加强NADH供给和阻断乙酸产生支路可能是提高C.acetoacidophilum菌株产琥珀酸产量的有效途径。
Lactate and succinate were produced by Corynebacterium acetoacidophilum from glucose under oxygen deprivation conditions. To construct knockout mutant, lactate dehydrogenase gene （ldh） of C. acetoacidophilum was deleted by double-crossover chromosome replacement with sacB gene. Comparing with the wild strain ATCC13870, ldhA-deficent mutant produced no lactate with glucose consumption rate decreased by 29.3%, while succinate and acetate concentrations were increased by 45.6% and 182%, respectively. Moreover, the NADH/NAD^＋ rate was less than 1 （about 0.7）, and the activities of phosphoenolpyruvate carboxylase and acetate kinase of the ldhA-deficent mutant were enhanced by 84% and 12 times, respectively. Our studies show that succinicate and acetate production pathways are strengthened by blocking lactate synthesis. It also suggests that improving NADH supply and eliminating acetate generation are alternative strategies to get high succinate-producer.
出处
《生物工程学报》
CAS
CSCD
北大核心
2014年第3期435-444,共10页
Chinese Journal of Biotechnology
基金
国家高技术研究发展计划(863计划)(No.2006AA020301-012)资助~~
关键词
嗜乙酰乙酸棒杆菌
ldhA-敲除菌
缺氧
琥珀酸
代谢途径
Corynebacterium acetoacidophilum, ldhA-deficent mutant, oxygen deprivation, succinic acid, metabolic
