Discovery and implementation of a novel pathway for n-butanol production via 2-oxoglutarate

作     者：Ferreira, Sofia Pereira, Rui Liu, Filipe Vilaca, Paulo Rocha, Isabel 

作者机构：Univ Minho CEB Ctr Biol Engn Campus Gualtar P-4710057 Braga Portugal SilicoLife Lda Rua Canastreiro 15 P-4715387 Braga Portugal Chalmers Univ Technol Dept Biol & Biol Engn S-41296 Gothenburg Sweden Argonne Natl Lab Math & Comp Sci Div 9700 S Cass Ave Argonne IL 60439 USA Univ Nova Lisboa ITQB NOVA Inst Tecnol Quim & Biol Antonio Xavier Oeiras Portugal 

出 版 物：《BIOTECHNOLOGY FOR BIOFUELS》 (生物燃料)

年 卷 期：2019年第12卷第1期

页      面：230-230页

核心收录：

学科分类：0820[工学-石油与天然气工程] 080703[工学-动力机械及工程] 08[工学] 0807[工学-动力工程及工程热物理] 0836[工学-生物工程] 

基　　金：Portuguese Foundation for Science and Technology (FCT) from MIT Portugal Program [PD/BD/52366/2013] European Regional Development Fund [POCI-01-0145-FEDER-006684, NORTE-01-0145-FEDER-000004] FCT/MCTES [ERA-IB-2/0002/2014] European Commission [H2020-LEIT-BIO-2015-1 686070-1] FEDER funds through COMPETE2020 Programa Operacional Competitividade e Internacionalizacao (POCI) [LISBOA010145FEDER007660] FCT Fundacao para a Ciencia e a Tecnologia UID/BIO/04469 Fundação para a Ciência e a Tecnologia [ERA-IB-2/0002/2014, PD/BD/52366/2013] Funding Source: FCT 

主　　题：n-Butanol E coli Metabolic engineering Enumeration algorithms 2-Oxoglutarate 

摘      要：Background One of the European Union directives indicates that 10% of all fuels must be bio-synthesized by 2020. In this regard, biobutanol-natively produced by clostridial strains-poses as a promising alternative biofuel. One possible approach to overcome the difficulties of the industrial exploration of the native producers is the expression of more suitable pathways in robust microorganisms such as Escherichia coli. The enumeration of novel pathways is a powerful tool, allowing to identify non-obvious combinations of enzymes to produce a target compound. Results This work describes the in silico driven design of E. coli strains able to produce butanol via 2-oxoglutarate by a novel pathway. This butanol pathway was generated by a hypergraph algorithm and selected from an initial set of 105,954 different routes by successively applying different filters, such as stoichiometric feasibility, size and novelty. The implementation of this pathway involved seven catalytic steps and required the insertion of nine heterologous genes from various sources in E. coli distributed in three plasmids. Expressing butanol genes in E. coli K12 and cultivation in High-Density Medium formulation seem to favor butanol accumulation via the 2-oxoglutarate pathway. The maximum butanol titer obtained was 85 +/- 1 mg L-1 by cultivating the cells in bioreactors. Conclusions In this work, we were able to successfully translate the computational analysis into in vivo applications, designing novel strains of E. coli able to produce n-butanol via an innovative pathway. Our results demonstrate that enumeration algorithms can broad the spectrum of butanol producing pathways. This validation encourages further research to other target compounds.