Metabolic engineering of Clostridium beijerinckii to improve glycerol metabolism and furfural tolerance

• Published in: Biotechnology for biofuels 2019-03, Vol.12 (1), p.50-50, Article 50
• Main Authors: Agu, Chidozie Victor, Ujor, Victor, Ezeji, Thaddeus Chukwuemeka
• Format: Article
• Language: English
• Subjects: Acetone; Additives; Alcohol; Bacteria; Biodiesel fuels; biomass; Butanol; C. beijerinckii; C. pasteurianum; Carbon; Cell growth; Clostridium; Clostridium beijerinckii; Clostridium pasteurianum; Dehydrogenases; Detoxification; Dihydroxyacetone; Engineering; Enzymes; Ethanol; Fermentation; Furfural; Fused protein; Gene expression; gene overexpression; Genes; Glycerol; Glycerol dehydrogenase; Glycerone kinase; Hydrolysates; Kinases; Lignocellulose; Metabolic engineering; Metabolism; Microorganisms; Monosaccharides; NAD; NADP (coenzyme); Overexpression; plasmids; Production management; Proteins; Raw materials; Regeneration; Substrates; Toxicity; Utilization
• ISSN: 1754-6834; 1754-6834
• DOI: 10.1186/s13068-019-1388-9

Inefficient utilization of glycerol by ( ) is a major impediment to adopting glycerol metabolism as a strategy for increasing NAD(P)H regeneration, which would in turn, alleviate the toxicity of lignocellulose-derived microbial inhibitory compounds (LDMICs, e.g., furfural), and improve the fermentation of lignocellulosic biomass hydrolysates (LBH) to butanol. To address this problem, we employed a metabolic engineering strategy to enhance glycerol utilization by . By overexpressing two glycerol dehydrogenase (Gldh) genes ( and ) from the glycerol hyper-utilizing ( ) as a fused protein in , we achieved approximately 43% increase in glycerol consumption, when compared to the plasmid control. Further, _ +  achieved a 59% increase in growth, while butanol and acetone-butanol-ethanol (ABE) concentrations and productivities increased 14.0%, 17.3%, and 55.6%, respectively, relative to the control. Co-expression of +  and + dihydroxyacetone kinase ( ) resulted in significant payoffs in cell growth and ABE production compared to expression of one Gldh. In the presence of 4-6 g/L furfural, increased glycerol consumption by the +  strain increased cell growth (> 50%), the rate of furfural detoxification (up to 68%), and ABE production (up to 40%), relative to the plasmid control. Likewise, over-expression of [( +  ) ] improved butanol and ABE production by 70% and 50%, respectively, in the presence of 5 and 6 g/L furfural relative to the plasmid control. Overexpression of and in significantly enhanced glycerol utilization, ABE production, and furfural tolerance by . Future research will address the inability of recombinant to metabolize glycerol as a sole substrate.