摘要
研究葡萄糖-6-磷酸脱氢酶基因g6pd过量表达对Candida tropicalis木糖醇生物合成代谢的影响。克隆Candida tropicalis CT16的g6pd基因,并将其与表达载体pYES-pgk重组连接,构建重组载体pYES-pgk-g6pd,LiAc/ssDNA/PEG方法转化导入C.tropicalis CT16,筛选阳性转化子,实现g6pd基因的过量表达。结果表明:发酵62 h,阳性转化子C.tropicalis SYG5的葡萄糖-6-磷酸脱氢酶活力提高了300%,发酵液中木糖醇质量浓度达到79.90 g/L,较野生型对照菌株的木糖醇产量提高了12.41%,木糖醇产率提高了44.94%。因此,葡萄糖-6-磷酸脱氢酶在C.tropicalis木糖醇的合成代谢途径中发挥重要作用,增强g6pd基因的表达,可以明显提高菌体NADPH供应量和还原力,有利于木糖醇的生物合成。
The influence of overexpression of glucose 6-phosphate dehydrogenase (G6PDH) gene,g6pd,on xylitol biosynthesis in Candida tropicalis was investigated.The gene g6pd was cloned from Candida tropicalis CT16 and inserted into a yeast expression vector pYES-pgk,generating a recombinant expression vector pYES-pgk-g6pd,which was then introduced into C.tropicalis CT16 by the LiAc/ssDNA/PEG transformation method,resulting in over-expression of the g6pd gene.The fermentation results showed that the G6PDH activity was increased by 300%,and a maximum xylitol yield of 79.90 g/L was achieved in the recombinant strain SYG5 harboring pYES-pgk-g6pd after 62 h of fermentation.Compared to the wild type strain C.tropicalis CT16,the yield and productive rate of xylitol in strain SYG5 were increased by 11.30% and 44.91%,respectively.These results indicate that increasing the expression level of the gene g6pd significantly enhances xylitol production and that the G6PDH plays a key role in the biosynthesis pathway of xylitol in C.tropicalis.
出处
《食品科学》
EI
CAS
CSCD
北大核心
2014年第7期102-106,共5页
Food Science
基金
国家高技术研究发展计划(863计划)项目(2012AA021502)
北京市自然科学基金项目(5122008)
教育部科学技术研究重点项目(211101)
北京市教委科技面上项目(KM201110011001)