Pichia stipitis木糖醇脱氢酶基因XYL2在酿酒酵母中的表达

通过PCR方法克隆得到树干毕赤氏酵母木糖醇脱氢酶(XDH)基因XYL2.将该基因连入酵母表达载体pYX212的强启动子磷酸丙糖异构酶(TPI)启动子下,得到融合表达载体pYX-XYL2.通过电转化方法将pYX-XYL2转入酿酒酵母Saccharomyces cerevisiae W303-1A中,酶活测定表明在酿酒酵母中树干毕赤氏酵母木糖醇脱氢酶基因XYL2得到活性表达,酿酒酵母转化子粗酶液中木糖醇脱氢酶比活为每毫克蛋白0.6 U左右,约为供体菌的2.4倍.与基因供体菌不同,木糖醇脱氢酶基因在酿酒酵母中表达不需木糖诱导,为组成型表达.

Expression of Pichia stipitis Xylitol Dehydrogenase Gene XYL2 in Saccharomyces cerevisiae

Pichia stipitis Xylitol dehydrogenase (XDH) Gene XYL2 was amplified by PCR. The XYL2 gene were placed under the triose phosphate isomerase (TPI) promoter of yeast vector pYX212 to produce the fusion expression vector pYX-XYL2. pYX-XYL2 was transformed into Saccharomyces cerevisiae W303-1A by electroporation. Transformants of S. cerevisiae containing XYL2 of P. stipitis synthesize an active XDH. Specif ic XDH activities of two transformants were 0. 59 U/mg protein and 0. 66 U/mg protein respectively, which were more than 2. 4 times of that of P. stipits CBS 5773 stain. Differently from P, stipits CBS 5773, the XYL2 in S. cerevisiae transformants expressed con-stitutively and did not need xylose for induction.