Improving formaldehyde consumption drives methanol assimilation in engineered E. coli

Woolston, Benjamin M.; King, Jason R.; Reiter, Michael; Van Hove, Bob; Stephanopoulos, Gregory

doi:10.1038/s41467-018-04795-4

Title: Improving formaldehyde consumption drives methanol assimilation in engineered E. coli

Journal Article · Tue Jun 19 00:00:00 EDT 2018 · Nature Communications

DOI:https://doi.org/10.1038/s41467-018-04795-4· OSTI ID:1454342

Woolston, Benjamin M.; King, Jason R.; Reiter, Michael; Van Hove, Bob; Stephanopoulos, Gregory

Abstract Due to volatile sugar prices, the food vs fuel debate, and recent increases in the supply of natural gas, methanol has emerged as a promising feedstock for the bio-based economy. However, attempts to engineer Escherichia coli to metabolize methanol have achieved limited success. Here, we provide a rigorous systematic analysis of several potential pathway bottlenecks. We show that regeneration of ribulose 5-phosphate in E. coli is insufficient to sustain methanol assimilation, and overcome this by activating the sedoheptulose bisphosphatase variant of the ribulose monophosphate pathway. By leveraging the kinetic isotope effect associated with deuterated methanol as a chemical probe, we further demonstrate that under these conditions overall pathway flux is kinetically limited by methanol dehydrogenase. Finally, we identify NADH as a potent kinetic inhibitor of this enzyme. These results provide direction for future engineering strategies to improve methanol utilization, and underscore the value of chemical biology methodologies in metabolic engineering.

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Research Organization:: Massachusetts Inst. of Technology (MIT), Cambridge, MA (United States)

Sponsoring Organization:: USDOE Advanced Research Projects Agency - Energy (ARPA-E); National Science Foundation (NSF); Fund for Scientific Research-Flanders (FWO)

Grant/Contract Number:: AR0000433; 1122374

OSTI ID:: 1454342

Alternate ID(s):: OSTI ID: 1904830

Journal Information:: Nature Communications, Journal Name: Nature Communications Vol. 9 Journal Issue: 1; ISSN 2041-1723

Publisher:: Nature Publishing GroupCopyright Statement

Country of Publication:: United Kingdom

Language:: English

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Cited by: 62 works

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