Journal
METABOLIC ENGINEERING
Volume 49, Issue -, Pages 257-266Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.ymben.2018.08.010
Keywords
Metabolic engineering; Synthetic biology; Methanol; Biofuel; C1 metabolism; Escherichia coli; 1-butanol; Ethanol
Categories
Funding
- Reducing Emissions using Methanotrophic Organisms for Transportation Energy (REMOTE) program of the Advanced Research Projects Agency-Energy [DE-AR0000430]
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Methanol is a potentially attractive substrate for bioproduction of chemicals because of the abundance of natural gas and biogas-derived methane. To move towards utilizing methanol as a sole carbon source, here we engineer an Escherichia coli strain to couple methanol utilization with growth on five-carbon (C5) sugars. By deleting essential genes in the pentose phosphate pathway for pentose utilization and expressing heterologous enzymes from the ribulose-monophosphate (RuMP) pathway, we constructed a strain that cannot grow on xylose or ribose minimal media unless methanol is utilized, creating a phenotype termed synthetic methanol auxotrophy. Our best strains were able to utilize methanol for growth at a rate of 0.17 +/- 0.006 (h(-1)) with methanol and xylose co-assimilation at a molar ratio of approximately 1: 1. Genome sequencing and reversion of mutations indicated that mutations on genes encoding for adenylate cyclase (cyaA) and the formaldehyde detoxification operon (frmRAB) were necessary for the growth phenotype. The methanol auxotrophic strain was further engineered to produce ethanol or 1-butanol to final titers of 4.6 g/L and 2.0 g/L, respectively. C-13 tracing showed that 43% and 71% of ethanol and 1-butanol produced had labeled carbon derived from methanol, respectively.
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