With growing concern regarding the depletion of fossil fuels, it has become increasingly important to design syntheses that are both sustainable and renewable. Recent advances in synthetic biology and metabolic engineering techniques have enabled the biosynthesis of non-natural metabolites in genetically engineered Escherichia coli (E. coli). One application of these strategies is the biosynthesis of citramalate, a chemical precursor to isoprene. The biosynthesis of isoprene using fermentation offers a more green and cost-effective alternative to harsher methods used in chemical processes. In this work we examine citramalate production in both wild type and knockout (ΔleuC and ΔleuD coding for 2-methylmalate hydrolyase) E. coli strains expressing the citramalate synthase (CimA) enzyme. CimA works to redirect natural metabolism by combining pyruvate with acetyl-CoA to catalyze the synthesis of citramalate. Wild type and knockout E. coli strains accumulated approximately 8.5 g/L and 13.4 g/L of citramalate respectively during 48 hours of fermentation. This resulted in over 52% and 82% of the theoretical maximum yield respectively from glucose as the sole carbon source. The ~1.6 fold increase in citramalate production affirms the ability of ΔleuC and ΔleuD knockout strains to drive metabolic flux forward by decreasing feedback inhibition from L-isoleucine production thereby increasing product yield.

Bio-production of isoprene precursor using engineered E. coli

2019 Spring National Meeting

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