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Engineering the biological conversion of methanol to specialty chemicals in Escherichia coli

Whitaker, W. Brian, Jones, J. Andrew, Bennett, R. Kyle, Gonzalez, Jacqueline E., Vernacchio, Victoria R., Collins, Shannon M., Palmer, Michael A., Schmidt, Samuel, Antoniewicz, Maciek R., Koffas, Mattheos A., Papoutsakis, Eleftherios T.
Metabolic engineering 2017 v.39 pp. 49-59
Escherichia coli, Geobacillus stearothermophilus, bioengineering, biological production, biomass, enzyme kinetics, enzymes, fuels, glycolysis, malates, metabolites, methanol, naringenin, ribulose, succinic acid, tricarboxylic acid cycle
Methanol is an attractive substrate for biological production of chemicals and fuels. Engineering methylotrophic Escherichia coli as a platform organism for converting methanol to metabolites is desirable. Prior efforts to engineer methylotrophic E. coli were limited by methanol dehydrogenases (Mdhs) with unfavorable enzyme kinetics. We engineered E. coli to utilize methanol using a superior NAD-dependent Mdh from Bacillus stearothermophilus and ribulose monophosphate (RuMP) pathway enzymes from B. methanolicus. Using ¹³C-labeling, we demonstrate this E. coli strain converts methanol into biomass components. For example, the key TCA cycle intermediates, succinate and malate, exhibit labeling up to 39%, while the lower glycolytic intermediate, 3-phosphoglycerate, up to 53%. Multiple carbons are labeled for each compound, demonstrating a cycling RuMP pathway for methanol assimilation to support growth. By incorporating the pathway to synthesize the flavanone naringenin, we demonstrate the first example of in vivo conversion of methanol into a specialty chemical in E. coli.