大肠杆菌混菌发酵生产咖啡醇糖苷  

Production of caffeyl alcohol glucosides by mixed fermentation with different Escherichia coli recombinants

作　　者：王帅 庄以彬[2] 刘浩[1] 毕慧萍[2] 刘涛[2] WANG Shuai;ZHUANG Yibin;LIU Hao;BI Huiping;LIU Tao(College of Engineering,Tianjin University of Science and Technology,Tianjin 300457,China;Tianjin Institute of Industrial Biotechnology,Chinese Academy of Sciences,Tianjin 300308,China)

机构地区：[1]天津科技大学生物工程学院,天津300457 [2]中国科学院天津工业生物技术研究所,天津300308

出　　处：《食品与发酵工业》2020年第23期7-13,共7页Food and Fermentation Industries

基　　金：国家自然科学基金项目(31970065)。

摘　　要：利用大肠杆菌混菌发酵生物合成咖啡醇糖苷。通过在酪氨酸高产大肠杆菌BTAL中表达异源基因酪氨酸解氨酶(tyrosine ammonia-lyase from Rhodotorula glutinis,RgTAL),对羟基肉桂酰辅酶A连接酶(hydroxycinnamate:CoA ligase from Petroselinum crispum,Pc4CL)和肉桂酰辅酶A还原酶(cinnamyl-CoA reductase from Arabidopsis thaliana,AtCCR),与大肠杆菌BL21(DE3)中表达对羟基苯乙酸3-羟化酶HpaBC和糖基转移酶(UGT73C5 from A.thaliana,AtUGT73C5)混合培养发酵生产了3种咖啡醇葡萄糖苷,其中2种为非天然化合物。优化共培养比例至3∶1,咖啡醇-4-O-葡萄糖苷的最高产量达到(141.63±3.42)mg/L。这是首次微生物异源合成咖啡醇糖苷的报道,该工作扩大了咖啡醇糖苷的结构多样性,为后续的构效分析提供了新的候选化合物,有望得到具有较好生物活性的咖啡醇糖苷化合物。Escherichia coli co-culture was designed for biosynthesis of caffeyl alcohol glucosides from glucose.Three caffeyl alcohol monoglucosides,namely caffeyl alcohol-3-O-glucoside,caffeyl alcohol-9-O-glucoside and caffeyl alcohol-4-O-glucoside,were produced by co-culturing two recombinant E.coli strains.A biosynthetic pathway of p-coumaryl alcohol was constructed in the high-tyrosine-producing E.coli BTAL-CAD01 by expressing tyrosine ammonialyase,hydroxycinnamate:CoA ligase(4CL),and cinnamyl-CoA reductase from Rhodotorula glutinis,Petroselinum crispum,and Arabidopsis thaliana,respectively,together with endogenous alcohol dehydrogenases or aldo-keto reductases.Then,4-hydroxyphenlacetate 3-hydroxylase(HpaBC)from E.coli and glucosyltransferase UGT73C5 from A.thaliana were co-expressed in wild-type E.coli BL21(DE3),resulting in a recombinant strain BWT-CAD01.Caffeyl alcohol monoglucosides were successfully produced by co-cultivating the recombinant strains BTAL-CAD01 and BWT-CAD01.The highest titer of caffeyl alcohol-4-O-glucoside reached(141.63±3.42)mg/L,with optimized inoculation ratio of 3∶1.To our knowledge,this is the first report on heterologous biosynthesis of caffeyl alcohol glucosides,both caffeyl alcohol-3-O-glucoside and caffeyl alcohol-9-O-glucoside are unnatural compounds.This work expands the structural diversity of caffeyl alcohol glucosides,which makes it possible the discovery of new analogues with improved pharmaceutical properties.

关 键 词：咖啡醇糖苷 生物合成 大肠杆菌 混菌培养 发酵生产 

分 类 号：TS201.3[轻工技术与工程—食品科学]