Improving formaldehyde consumption drives methanol assimilation in engineered E. coli
Abstract
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.
- Authors:
- Publication Date:
- Research Org.:
- Massachusetts Inst. of Technology (MIT), Cambridge, MA (United States)
- Sponsoring Org.:
- USDOE Advanced Research Projects Agency - Energy (ARPA-E); National Science Foundation (NSF); Fund for Scientific Research-Flanders (FWO)
- OSTI Identifier:
- 1454342
- Alternate Identifier(s):
- OSTI ID: 1904830
- Grant/Contract Number:
- AR0000433; 1122374
- Resource Type:
- Published Article
- Journal Name:
- Nature Communications
- Additional Journal Information:
- Journal Name: Nature Communications Journal Volume: 9 Journal Issue: 1; Journal ID: ISSN 2041-1723
- Publisher:
- Nature Publishing Group
- Country of Publication:
- United Kingdom
- Language:
- English
- Subject:
- 59 BASIC BIOLOGICAL SCIENCES; chemical genetics; metabolic engineering; metabolomics
Citation Formats
Woolston, Benjamin M., King, Jason R., Reiter, Michael, Van Hove, Bob, and Stephanopoulos, Gregory. Improving formaldehyde consumption drives methanol assimilation in engineered E. coli. United Kingdom: N. p., 2018.
Web. doi:10.1038/s41467-018-04795-4.
Woolston, Benjamin M., King, Jason R., Reiter, Michael, Van Hove, Bob, & Stephanopoulos, Gregory. Improving formaldehyde consumption drives methanol assimilation in engineered E. coli. United Kingdom. https://doi.org/10.1038/s41467-018-04795-4
Woolston, Benjamin M., King, Jason R., Reiter, Michael, Van Hove, Bob, and Stephanopoulos, Gregory. Tue .
"Improving formaldehyde consumption drives methanol assimilation in engineered E. coli". United Kingdom. https://doi.org/10.1038/s41467-018-04795-4.
@article{osti_1454342,
title = {Improving formaldehyde consumption drives methanol assimilation in engineered E. coli},
author = {Woolston, Benjamin M. and King, Jason R. and Reiter, Michael and Van Hove, Bob and Stephanopoulos, Gregory},
abstractNote = {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.},
doi = {10.1038/s41467-018-04795-4},
journal = {Nature Communications},
number = 1,
volume = 9,
place = {United Kingdom},
year = {Tue Jun 19 00:00:00 EDT 2018},
month = {Tue Jun 19 00:00:00 EDT 2018}
}
https://doi.org/10.1038/s41467-018-04795-4
Web of Science
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