Metabolic engineering of Clostridium cellulolyticum for the production of n-butanol from crystalline cellulose

Microbial Cell Factories

Research

Andreas Heinz Wilhelm Jentges¹  Stefan Marcus Gaida¹  Benedikt Engels¹  Andrea Liedtke¹  Stefan Jennewein¹ 

[1] Department of Industrial Biotechnology, Fraunhofer Institute for Molecular Biology and Applied Ecology, Forckenbeckstrasse 6, 52074, Aachen, Germany;

关键词: Metabolic engineering;    Clostridium cellulolyticum;    Biofuels;    Butanol;    Clostridia;   

DOI  :  10.1186/s12934-015-0406-2

 received in 2015-08-11, accepted in 2015-12-19,  发布年份 2016

来源: Springer

【 摘 要 】

BackgroundSustainable alternatives for the production of fuels and chemicals are needed to reduce our dependency on fossil resources and to avoid the negative impact of their excessive use on the global climate. Lignocellulosic feedstock from agricultural residues, energy crops and municipal solid waste provides an abundant and carbon-neutral alternative, but it is recalcitrant towards microbial degradation and must therefore undergo extensive pretreatment to release the monomeric sugar units used by biofuel-producing microbes. These pretreatment steps can be reduced by using microbes such as Clostridium cellulolyticum that naturally digest lignocellulose, but this limits the range of biofuels that can be produced. We therefore developed a metabolic engineering approach in C. cellulolyticum to expand its natural product spectrum and to fine tune the engineered metabolic pathways.ResultsHere we report the metabolic engineering of C. cellulolyticum to produce n-butanol, a next-generation biofuel and important chemical feedstock, directly from crystalline cellulose. We introduced the CoA-dependent pathway for n-butanol synthesis from C. acetobutylicum and measured the expression of functional enzymes (using targeted proteomics) and the abundance of metabolic intermediates (by LC-MS/MS) to identify potential bottlenecks in the n-butanol biosynthesis pathway. We achieved yields of 40 and 120 mg/L n-butanol from cellobiose and crystalline cellulose, respectively, after cultivating the bacteria for 6 and 20 days.ConclusionThe analysis of enzyme activities and key intracellular metabolites provides a robust framework to determine the metabolic flux through heterologous pathways in C. cellulolyticum, allowing further improvements by fine tuning individual steps to improve the yields of n-butanol.

【 授权许可】

CC BY   
© Gaida et al. 2016