酿酒酵母工业菌株中XI木糖代谢途径的建立

根据代谢工程原理,采取多拷贝整合策略,利用整合载体pYMIKP,将来自嗜热细菌Thermusthermophilus的木糖异构酶(XI)基因xylA和酿酒酵母(Saccharomycescerevisiae)自身的木酮糖激酶(XK)基因XKS1,插入酿酒酵母工业菌株NAN-27的染色体中,得到工程菌株NAN-114。酶活测定结果显示,NAN-114中XI和XK的活性均高于出发菌株NAN-27,表明外源蛋白在酿酒酵母工业菌株中得到活性表达。对木糖、葡萄糖共发酵摇瓶实验结果表明,工程菌NAN-114消耗木糖4.6g/L,产生乙醇6.9g/L,较出发菌株分别提高了43.8%和9.5%。首次在酿酒酵母工业菌株中建立了XI路径的木糖代谢途径。

Construction of Industrial Saccharomyces cerevisiae Expressing Xylose-Metabolizing Genes in XI Pathway

In order to develop an industrial strain of Saccharomyces cerevisiae that can utilize the xylose to produce ethanol, an integrating plasmid pYMIK-xy114 containing the xylA gene from Thermus thermophilus, encoding xylose isomerase (XI), and XKS1 gene from S. cerevisiae, encoding xylulokinase (XK) was constructed. The integrating plasmid pYMIK-xy114 was transformed into a S. cerevisiae industrial strain NAN-27 producing the recombinant strain NAN-114. Upon transformation, multiple copies of the xylose-utilizing genes were integrated into the genome rDNA locus of S. cerevisiae. The results of enzymes assays showed that the XI and XK activity of NAN-114 was higher than that of parent strain. The resulting recombinant strain could co-ferment glucose and xylose to ethanol under oxygen-limited condition. The NAN-114 consumed 4.6g/L xylose and produced 6.9g/L ethanol, which were 43.8% and 9.5% higher than the parent strain NAN-27 respectively.