乙醇和木糖醇联产酿酒酵母菌株的代谢优化  

Metabolic optimization of an ethanol and xylitol-coproducing Saccharomyces cerevisiae strain

作　　者：王芯雨 杨白雪[1,2] 谢采芸 张全 缪晡 汤岳琴 WANG Xinyu;YANG Baixue;XIE Caiyun;ZHANG Quan;MIAO Bu;TANG Yueqin(Environmental Biotechnology Research Center,College of Architecture and Environment,Sichuan University,Chengdu 610065,China;Sichuan Environmental Protection Key Laboratory of Organic Wastes Valorization,Chengdu 610065,China;Sinopec(Dalian)Research Institute of Petroleum and Petrochemicals Co.,Ltd.,Dalian 116045,China;Sinopec Shanghai Engineering Co.,Ltd.,Shanghai 200120,China)

机构地区：[1]四川大学建筑与环境学院环境生物技术研究中心,成都610065 [2]四川省环境保护有机废弃物资源化利用重点实验室,成都610065 [3]中石化(大连)石油化工研究院有限公司,大连116045 [4]中石化上海工程有限公司,上海200120

出　　处：《应用与环境生物学报》2024年第3期586-592,共7页Chinese Journal of Applied and Environmental Biology

基　　金：国家重点研发计划重点专项项目(2022YFE0108500)资助。

摘　　要：构建性能优良的发酵葡萄糖和木糖同步生产乙醇和木糖醇的工业酿酒酵母菌株是实现秸秆等木质纤维素原料燃料乙醇和木糖醇同步生产的基础.以前期工作构建的一株具有良好木糖醇积累能力的工程菌株CK17为基础,利用CRISPR/Cas9基因编辑技术,研究菌株内源己糖转运蛋白定点突变、水甘油通道蛋白编码基因FPS1高表达、木糖还原基因拷贝数增加等对菌株在批次发酵条件下代谢木糖积累木糖醇的影响.结果表明,定点突变己糖转运蛋白Hxt3p和Hxt7p获得了工程菌株CM3和CM7,其木糖利用速率分别提升了92.1%和69.0%.将CM3的FPS1基因的启动子替换成TDH3基因启动子获得了工程菌株CM3FTD,其木糖利用速率比CM3提升了32.6%.增加CM3FTD菌株的木糖还原酶基因的拷贝数获得了菌株TX,其木糖利用速率比CM3FTD提升了17.2%.利用秸秆预处理物料进行同步糖化发酵,菌株TX发酵性能较出发菌株CK17得到显著提升,木糖代谢速率提升56.4%,木糖醇浓度和乙醇浓度分别达到57.6 g/kg和48.5g/kg,木糖醇收率接近理论收率,乙醇收率达到0.43g/g.本研究构建的工程菌株为利用秸秆原料同步生产乙醇和木糖醇提供了优良的菌株来源.Industrial Saccharomyces cerevisiae strains with excellent capacity for coproducing ethanol and xylitol from glucose and xylose are necessary for the coproduction of fuel ethanol and xylitol from lignocellulosic feedstocks such as agricultural straw.In this study,using the xylitol-producing strain CK17 as host,the effects of point mutations in the endogenous hexose transporter,elevated expression of the glycerol transporter channel Fps1p,and an increased copy number of xylose reductase-encoding genes on xylitol production were investigated using CRISPR/Cas9 gene editing technology.Point mutations in the hexose transporters Hxt3p and Hxt7p enhanced xylose consumption by 92.1%(strain CM3)and 69.0%(strain CM7),respectively.Replacing the promoter of the FPS1 gene of CM3 with the promoter of the TDH3 gene yielded strain CM3FTD,which showed a 32.6%enhancement over CM3 in its xylose consumption rate.Strain TX was obtained by increasing the copy number of the xylose reductase gene in strain CM3FTD.Its xylose consumption rate was enhanced by 17.2%over that of CM3FTD.When using pretreated straw for simultaneous saccharification and fermentation,strain TX showed significantly improved fermentation capacity compared with strain CK17,and its xylose consumption rate was enhanced by 56.4%.Cumulative xylose and ethanol reached 57.6 g/kg and 48.5 g/kg,respectively.Xylitol yield reached its theoretical maximum,and ethanol yield was 0.43 g/g-glucose.By mutating the hexose transporters and increasing Fps1p expression and xylose reductase gene copy number,S.cerevisiae strain TX with excellent ethanol-and xylitol-coproducing capacity was obtained.This strain exhibits good potential for coproduction of ethanol and xylitol from agricultural straw feedstocks.

关 键 词：酿酒酵母 木糖醇 纤维素燃料乙醇 秸秆 

分 类 号：TQ920.1[轻工技术与工程—发酵工程] TQ223.122