Reconstruction the feedback regulation of amino acid metabolism to develop a non-auxotrophic l-threonine producing Corynebacterium glutamicum  

作　　者：Jianhang Liu Jiao Liu Jiajun Li Xiaojia Zhao Guannan Sun Qianqian Qiao Tuo Shi Bin Che Jiuzhou Chen Qianqian Zhuang Yu Wang Jibin Sun Deqiang Zhu Ping Zheng 

机构地区：[1]State Key Laboratory of Biobased Material and Green Papermaking,Qilu University of Technology,Shandong Academy of Sciences,Jinan 250353,China [2]Key Laboratory of Engineering Biology for Low-carbon Manufacturing,Tianjin Institute of Industrial Biotechnology,Chinese Academy of Sciences,Tianjin 300308,China [3]National Center of Technology Innovation for Synthetic Biology,Tianjin 300308,China [4]Shandong Provincial Key Laboratory of Microbial Engineering,School of Bioengineering,Qilu University of Technology,Shandong Academy of Sciences,Jinan 250353,China [5]Shandong University of Traditional Chinese Medicine,Jinan 250355,China

出　　处：《Bioresources and Bioprocessing》2024年第1期599-609,共11页生物资源与生物加工(英文)

基　　金：funded by the National Key Research and Development Program of China(2021YFC2100900);the Tianjin Synthetic Biotechnology Innovation Capacity Improvement Project(TSBICIP-CXRC-058);the National Natural Science Foundation of China(32270101);the Key R&D Program of Shandong Province(2021CXGC010602);the Youth Innovation Promotion Association of Chinese Academy of Sciences(2021177)。

摘　　要：l-Threonine is an important feed additive with the third largest market size among the amino acids produced by microbial fermentation.The GRAS(generally regarded as safe)industrial workhorse Corynebacterium glutamicum is an attractive chassis for l-threonine production.However,the present l-threonine production in C.glutamicum cannot meet the requirement of industrialization due to the relatively low production level of l-threonine and the accumulation of large amounts of by-products(such as l-lysine,l-isoleucine,and glycine).Herein,to enhance the l-threonine biosynthesis in C.glutamicum,releasing the aspartate kinase(LysC)and homoserine dehydrogenase(Hom)from feedback inhibition by l-lysine and l-threonine,respectively,and overexpressing four flux-control genes were performed.Next,to reduce the formation of by-products l-lysine and l-isoleucine without the cause of an auxotrophic phenotype,the feedback regulation of dihydrodipicolinate synthase(DapA)and threonine dehydratase(IlvA)was strengthened by replacing the native enzymes with heterologous analogues with more sensitive feedback inhibition by l-lysine and l-isoleucine,respectively.The resulting strain maintained the capability of synthesizing enough amounts of l-lysine and l-isoleucine for cell biomass formation but exhibited almost no extracellular accumulation of these two amino acids.To further enhance l-threonine production and reduce the by-product glycine,l-threonine exporter and homoserine kinase were overexpressed.Finally,the rationally engineered non-auxotrophic strain ZcglT9 produced 67.63 g/L(17.2%higher)l-threonine with a productivity of 1.20 g/L/h(108.0%higher)in fed-batch fermentation,along with significantly reduced by-product accumulation,representing the record for l-threonine production in C.glutamicum.In this study,we developed a strategy of reconstructing the feedback regulation of amino acid metabolism and successfully applied this strategy to de novo construct a non-auxotrophic l-threonine producing C.glutamicum.The main end by-product

关 键 词：L-THREONINE Corynebacterium glutamicum BY-PRODUCT Allosteric regulation Transport engineering 

分 类 号：O62[理学—有机化学]