(288a) A New-to-Nature Metabolic Pathway for Methanol and Formaldehyde Assimilation in Escherichia coli | AIChE

(288a) A New-to-Nature Metabolic Pathway for Methanol and Formaldehyde Assimilation in Escherichia coli

Authors 

Lee, S. H. - Presenter, Rice University
Chou, A., University of South Florida
Clomburg, J. M., University of South Florida
Gonzalez, R., University of South Florida
Zhu, F., University of South Florida
One-carbon (C1) bioconversion has been recognized as a potential solution to the global decarbonization goal and the mitigation of global warming and climate change. Among various C1 compounds, methanol has been a widely sought feedstock for bioconversion because it is a liquid at ambient conditions and has a high energy content compared to more oxidized forms. Formaldehyde, which is the product of methanol oxidation catalyzed by methanol dehydrogenase, is a common node for many pathways involved in C1 metabolism. While formaldehyde has the same oxidation state as sugars, which is advantageous to achieve redox balance during fermentative metabolism, its high toxicity makes it a challenging feedstocks for bioconversions.

Here, we present a new-to-nature metabolic pathway that efficiently utilize C1 substrates such as methanol and formaldehyde through C1-C1 condensation reactions catalyzed by the enzyme 2-hydroxyacyl-CoA lyase (HACL). Unlike other native C1 bioconversion pathways such as RuMP cycle and Serine pathway, the HACL-based pathway is orthogonal to the central metabolism. It is catalyzed by only 2 essential enzymes (HACL and acyl-CoA reductase (ACR)) to a precursor metabolite glycolate, when started from formaldehyde. Glycolate can be used as substrate for cell growth or its precursor CoA-thioester (glycolyl-CoA) converted orthogonally to longer-chain products via multiple iteration of formyl-CoA elongation reactions. We have demonstrated the orthogonality of the pathway by splitting the C1-bioconversion pathway and the cell growth in a two-strain, E. coli-E. coli co-culture system. The producer strain harbors expression vectors for glycolate producing enzymes but is deficient in the glycolate-utilization pathway. The sensor strain is wildtype MG1655 able to grow on glycolate as sole carbon source, but unable to utilize C1 substrates. The sensor strain is able to grow on the glycolate produced by the producer strain in a co-culture system when methanol, formaldehyde and the mixture of formaldehyde and formate were provided as the only carbon sources. We also demonstrate the ability of HACL-based pathways to produce diverse C2 and C3 products, including but not limited to glycolaldehyde, ethylene glycol, acetate, ethanol and glycerate.