调控乙酰CoA合成酶表达促进乙酸利用及GlcNAc合成  

Modulation of Acetyl-CoA Synthetase Expression Promoted Acetate Utilization and N-Acetylglucosamine Production in Bacillus subtilis

作　　者：马文龙 刘延峰 吕雪芹 李江华[1,2] 刘龙[1,2] 堵国成[1,2] MA Wenlong;LIU Yanfeng;LYU Xueqin;LI Jianghua;LIU Long;DU Guocheng(Key Laboratory of Industrial Biotechnology,Ministry of Education,Jiangnan University,Wuxi 214122,China;School of Biotechnology,Jiangnan University,Wuxi 214122,China)

机构地区：[1]江南大学工业生物技术教育部重点实验室,江苏无锡214122 [2]江南大学生物工程学院,江苏无锡214122

出　　处：《食品与生物技术学报》2020年第5期16-22,共7页Journal of Food Science and Biotechnology

基　　金：国家自然科学基金项目(31622001,31671845,21676119,31600068);江苏省自然科学基金项目(BK20160176)。

摘　　要：N-乙酰氨基葡萄糖(GlcNAc)是一种重要的功能单糖。在利用枯草芽孢杆菌发酵产GlcNAc的过程中,乙酸的积累严重抑制了细胞生长和GlcNAc合成。为了获得高效利用乙酸生产GlcNAc的微生物细胞工厂,作者通过突变乙酰CoA(Ac-CoA)合成酶编码基因acsA 5′UTR区域内全局转录调控因子CodY结合序列,调控acsA转录水平,进一步突变AcsA乙酰化调控位点,解除乙酰化修饰,促进乙酸转化为Ac-CoA,最终乙酸积累量由6.3 g/L减少到3.6 g/L,同时细胞干重(DCW)由2.7 g/L增加到5.3 g/L,GlcNAc产量由1.9 g/L提高到5.0 g/L,为进一步获得高产GlcNAc细胞工厂提供了基础。N-acetyl-D-glucosamine(GlcNAc) is an important functional monosaccharide. In the production of GlcNAc using recombinant Bacillus subtillis(B. subtilis), accumulation of acetate inhibites cell growth and GlcNAc production, which also decreases GlcNAc yield. To construct a B.subtilis cell factory with efficient acetate utilization and GlcNAc production, expression of acetyl-CoA synthetase AcsA, which catalyzes the conversion of acetate to Ac-CoA, was modulated in this study. Specifically, the global transcriptional regulator CodY binding sites in the 5’ UTR region of acs A were mutated to promote transcription of acsA;then, enzyme AcsA was modified by deacetylation to improve its activity. Finally, the accumulated acetate during shake flask fermentation reduced from 6.3 to 3.6 g/L, dry cell weight increased from 2.7 to 5.3 g/L, and the GlcNAc yield was increased by 163% up to 5.0 g/L, which indicated that the developed strain offers improved properties for GlcNAc production.

关 键 词：枯草芽孢杆菌 乙酸 乙酰辅酶A合成酶 N-乙酰氨基葡萄糖 

分 类 号：Q815[生物学—生物工程]