Metabolic Engineering and Synthetic Biology

Metabolic engineering and synthetic biology are emerging disciplines that enable the design and engineering of biological systems for a wide range of applications. In this area, our laboratory has pioneered the engineering of glycerol fermentation and metabolism of waste fatty acids for the synthesis of fuels and chemicals and the development of a functional reversal of the β-oxidation cycle as an efficient platform for the synthesis of longer-chain (C ≥ 4) products.

The knowledge base created by fundamental studies of glycerol metabolism in our laboratory (see “Systems biology of metabolism“) has laid the foundation to establish glycerol fermentation as a new metabolic engineering platform for fuel and chemical production. We pioneered the engineering of bacteria to efficiently convert glycerol to fuels and chemicals such as succinate, ethanol, hydrogen, formate, D- and L-lactate, and 1,2-propanediol (Trends Biotechnol. 31: 20, 2013; Microb. Cell Fact. 12: 7, 2013; Appl. Bioch. Biotechnol. 166: 680, 2012; Biotechnol. Bioeng. 108: 86, 2011; Metab. Eng. 12: 409, 2010; Appl. Environ. Microbiol. 76: 4327, 2010; Biotechnol. Lett. 32: 405, 2010; Biotechnol. Bioeng. 103: 148, 2009; Metab. Eng. 10: 340, 2008; Curr. Opin. Biotechnol. 18: 213, 2007). We have also established a novel platform for the production of fuels and chemicals from fatty acid-rich feedstocks by engineering a respiro-fermentative metabolic mode that enables the efficient production of target products in combination with adequate catabolism of FAs. We engineered efficient synthesis of ethanol, butanol, acetate, acetone, isopropanol, succinate, and propionate from fatty acids in bacteria (Rev.-Syst. Biol. 5: 575, 2013; Appl. Environ. Microbiol. 76: 5067, 2010).

Advanced, higher-chain (C ≥ 4) fuels and chemicals are generated from short-chain, 2- or 3-C metabolic intermediates through pathways that require carbon-chain elongation. While our laboratory and others around the world have engineered native carbon-chain elongation pathways, such as the fatty acid biosynthesis pathway, to produce higher-chain molecules like methylketones (J. Industrial Microbiol. Biotechnol. 39: 1703, 2012), this pathway suffers from major energy constraints. Motivated in part by these limitations, we recently engineered a functional reversal of the β-oxidation cycle that can be used as a general platform for the synthesis of short-, medium- and long-chain products with structural and functional diversity (Metab. Eng. 28: 202, 2015; Appl. Environ. Microbiol. 81: 1406, 2015; J. Industrial Microbiol. Biotechnol. 42:465, 2015; ACS Synth. Biol. 1: 541, 2012; Nature 476: 355, 2011). Through a systems-level, quantitative assessment of the metabolic capabilities of the engineered reversal of the β-oxidation cycle, we demonstrated that product synthesis can be coupled to cell growth and achieved at high fluxes, titers and yields (Metab. Eng. 23: 100, 2014). The superior capabilities of the β-oxidation reversal, when compared to other pathways used for carbon-chain elongation, originate from its higher energetic efficiency, which is enabled by the use of acetyl-CoA as an extender unit. This engineered β-oxidation reversal is currently being exploited in our laboratory for the production of alcohols, alkanes, and omega-functionalized products.