启动子工程改造大肠杆菌K4生产果糖软骨素  

Promoter engineering of Escherichia coli K4 for production of fructosylated chondroitin

作　　者：吴秋林[1,2,3] 刘佳[1,2,3] 杨爱华[1,2,3] 刘立明[1,2,3] 

机构地区：[1]江南大学食品科学与技术国家重点实验室,无锡214122 [2]江南大学工业生物技术教育部重点实验室,无锡214122 [3]江南大学食品微生物制造工程实验室,无锡214122

出　　处：《生物加工过程》2014年第3期19-25,共7页Chinese Journal of Bioprocess Engineering

基　　金：中组部青年拔尖人才支持计划;国家重点基础研究发展计划(973计划)(2013CB733600);江苏省科技支撑计划(2012617);江苏省杰出青年基金(BK2012002)

摘　　要：为了进一步提高大肠杆菌K4发酵生产果糖软骨素(K4CPS)的产量,将合成基因簇region 3启动子(PR3)3'端非翻译区(UTR)进行缺失突变,研究其对突变菌株K4CPS合成的影响。研究表明:在ops序列(RfaH蛋白结合位点)存在时,PR3启动子强度和K4CPS产量与UTR的长度变化无关;但ops序列缺失时,UTR的延长可导致PR3启动子的强度和K4CPS产量均低于对照菌株;反之,UTR的缩短能显著提高PR3启动子的强度,进而使K4CPS产量比原菌增加了46%,达到751 mg/L。The capsular polysaccharide of Escherichia coli K4 was almost identical to chondroitin, a commercially valuable biopolymer.In order to further improve the productivity of K4CPS by E.coli K4, the modification of 3′ untranslated region（UTR） of the capsule gene cluster region 3 promoter（PR3） were investigated.The mutants with UTR deletion were screened and confirmed by PCR analysis and sequencing.The fermentation and quantitative real time RCR results suggested that the changes of UTR sequence as ops sequence remain did not affect the production of K4CPS and the transcription of PR3.Compared with the wild type,PR3 transcription level and the K4CPS yield of the insertion mutant with ops sequence deletion were decreased.Mutant strain was constructed by deleting both of UTR and ops sequence,however,PR3 transcription level was increased by 2-fold and the K4CPS yield reached 751 mg/L,with an increase of 46%.

关 键 词：硫酸软骨素 大肠杆菌K4 启动子工程 RED同源重组 

分 类 号：TQ923[轻工技术与工程—发酵工程]