Despite the potential of biotechnological processes for one-carbon (C1) bioconversion, efficient biocatalysts required for their implementation are yet to be developed. To address intrinsic limitations of native C1 biocatalysts, here we report that 2-hydroxyacyl CoA lyase (HACL), an enzyme involved in mammalian α-oxidation, catalyzes the ligation of carbonyl-containing molecules of different chain lengths with formyl-coenzyme A (CoA) to produce C1-elongated 2-hydroxyacyl-CoAs. We discovered and characterized a prokaryotic variant of HACL and identified critical residues for this newfound activity, including those supporting the hypothesized thiamine pyrophosphate-dependent acyloin condensation mechanism. The use of formyl-CoA as a C1 donor provides kinetic advantages and enables C1 bioconversion to multi-carbon products, demonstrated here by engineering an Escherichia coli whole-cell biotransformation system for the synthesis of glycolate and 2-hydroxyisobutyrate from formaldehyde and formaldehyde plus acetone, respectively. Our work establishes a new approach for C1 bioconversion and the potential for HACL-based pathways to support synthetic methylotrophy.
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We thank S. Garg for assistance with protein purification methods and S. Cheong for assistance with genetic methods. This work was partially supported by a grant from the US National Science Foundation (no. CBET-1605999).
A.C., J.M.C. and R.G. are co-inventors and assignees on a patent application (PCT/US2015/058121), which relates to the reported research. Correspondence and requests for materials should be addressed to email@example.com.
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Chou, A., Clomburg, J.M., Qian, S. et al. 2-Hydroxyacyl-CoA lyase catalyzes acyloin condensation for one-carbon bioconversion. Nat Chem Biol 15, 900–906 (2019). https://doi.org/10.1038/s41589-019-0328-0
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