Kogje Anushree, Ghosalkar Anand
Department of Technology, Savitribai Phule Pune University, Pune, Maharashtra, 411007, India.
Division of Praj Industries Limited, Praj-Matrix - R & D Centre, 402/403/1098, Urawade, Pune, Maharashtra, 412115, India.
3 Biotech. 2016 Dec;6(2):127. doi: 10.1007/s13205-016-0444-4. Epub 2016 Jun 7.
Xylitol production was compared in fed batch fermentation by Saccharomyces cerevisiae strains overexpressing xylose reductase (XR) genes from Candida tropicalis, Pichia stipitis, Neurospora crassa, and an endogenous gene GRE3. The gene encoding a xylose specific transporter (SUT1) from P. stipitis was cloned to improve xylose transport and fed batch fermentation was used with glucose as a cosubstrate to regenerate NADPH. Xylitol yield was near theoretical for all the strains in fed batch fermentation. The highest volumetric (0.28 gL h) and specific (34 mgg h) xylitol productivities were obtained by the strain overexpressing GRE3 gene, while the control strain showed 7.2 mgg h specific productivity. The recombinant strains carrying XR from C. tropicalis, P. stipitis, and N. crassa produced xylitol with lower specific productivity of 14.3, 6.8, and 6.3 mgg h, respectively, than GRE3 overexpressing strain. The glucose fed as cosubstrate was converted to biomass and ethanol, while xylose was only converted to xylitol. The efficiency of ethanol production was in the range of 38-45 % of the theoretical maximum for all the strains. Xylitol production from the non-detoxified corncob hemicellulosic hydrolysate by recombinant S. cerevisiae was reported for the first time. Xylitol productivity was found to be equivalent in the synthetic xylose as well as hemicellulosic hydrolysate-based media showing no inhibition on the S. cerevisiae due to the inhibitors present in the hydrolysate. A systematic evaluation of heterologous XRs and endogenous GRE3 genes was performed, and the strain overexpressing the endogenous GRE3 gene showed the best xylitol productivity.
通过酿酒酵母菌株在补料分批发酵中比较木糖醇的产量，这些菌株过表达来自热带假丝酵母、树干毕赤酵母、粗糙脉孢菌的木糖还原酶（XR）基因以及一个内源性基因GRE3。克隆了来自树干毕赤酵母的编码木糖特异性转运蛋白（SUT1）的基因以改善木糖转运，并使用葡萄糖作为共底物进行补料分批发酵以再生NADPH。在补料分批发酵中，所有菌株的木糖醇产量均接近理论值。过表达GRE3基因的菌株获得了最高的体积木糖醇生产率（0.28 g/L·h）和比木糖醇生产率（34 mg/g·h），而对照菌株的比生产率为7.2 mg/g·h。携带来自热带假丝酵母、树干毕赤酵母和粗糙脉孢菌的XR的重组菌株产生木糖醇的比生产率分别为14.3、6.8和6.3 mg/g·h，低于过表达GRE3基因的菌株。作为共底物添加的葡萄糖被转化为生物量和乙醇，而木糖仅被转化为木糖醇。所有菌株的乙醇生产效率在理论最大值的38 - 45%范围内。首次报道了重组酿酒酵母从未脱毒的玉米芯半纤维素水解物中生产木糖醇。发现在合成木糖以及基于半纤维素水解物的培养基中木糖醇生产率相当，表明水解物中存在的抑制剂对酿酒酵母没有抑制作用。对异源XR和内源性GRE3基因进行了系统评估，过表达内源性GRE3基因的菌株显示出最佳的木糖醇生产率。
