混合糖发酵重组酿酒酵母的菌株构建和菊芋秸秆同步糖化发酵研究

Construction of mixed-sugar fermenting recombinant Saccharomyces cerevisiae and ethanol production from Jerusalem artichoke stalk by simultaneous saccharification and fermentation

【目的】构建可用于纤维素乙醇高效生产的混合糖发酵重组酿酒酵母菌株,并利用菊芋秸秆为原料进行乙醇发酵。【方法】筛选在木糖中生长较好的酿酒酵母YB-2625作为宿主菌,构建木糖共代谢菌株YB-2625 CCX。进一步通过r DNA位点多拷贝整合的方式,以YB-2625 CCX为出发菌株构建木糖脱氢酶过表达菌株,并筛选得到优势菌株YB-73。采用同步糖化发酵策略研究YB-73的菊芋秸秆发酵性能。【结果】YB-73菌株以90 g/L葡萄糖和30 g/L木糖为碳源进行混合糖发酵,乙醇产量比出发菌株YB-2625 CCX提高了13.9%,副产物木糖醇产率由0.89 g/g降低至0.31 g/g,下降了64.6%。利用重组菌YB-73对菊芋秸秆进行同步糖化发酵,48 h最高乙醇浓度达到6.10%(体积比)。【结论】通过转入木糖代谢途径以及r DNA位点多拷贝整合过表达木糖脱氢酶基因可有效提高菌株木糖发酵性能,并用于菊芋秸秆的纤维素乙醇生产。这是首次报道利用重组酿酒酵母进行菊芋秸秆原料的纤维素乙醇发酵。

[Objective] This study aimed to construct highly efficient recombinant Saccharomyces cerevisiae strain for cellulosic bioethanol production from Jerusalem artichoke stalk（JAS）. [Methods] S. cerevisiae strain YB-2625 was selected as a host strain to construct xylose co-fermenting strain YB-2625 CCX, after which multicopies of xylitol dehydrogenase（XDH） encoding gene were integrated into the r DNA locus of YB-2625 CCX, and the most efficient strain named YB-73 was obtained. Finally, ethanol production from JAS was investigated by simultaneous saccharification and fermentation（SSF） using YB-73. [Results] YB-73 showed improved ethanol production by 13.9% compared with that of YB-2625 CCX and the xylitol yield of YB-73 was reduced to 0.31 g/g xylose from 0.89 g/g xylose by YB-2625 CCX when fermenting with 90 g/L glucose and 30 g/L xylose. Meanwhile, flocculation of YB-73 was observed in the presence of xylose, and the strain also showed high tolerance towards 5 g/L acetic acid and high temperature. The highest ethanol titer of 6.10%（V/V） was achieved from JAS in the process of SSF using YB-73. [Conclusion] Combination of host selection, introduction of xylose-consuming pathway and multi-copy overexpression of XDH in r DNA locus is a rational strategy to improve cellulosic bioethanol production performance of S. cerevisiae using JAS. This is the first report using recombinant S. cerevisiae to produce cellulosic ethanol from JAS.