Engineering the biological conversion of methanol to specialty chemicals in Escherichia coli

W Brian Whitaker; J Andrew Jones; R Kyle Bennett; Jacqueline E Gonzalez; Victoria R Vernacchio; Shannon M Collins; Michael A Palmer; Samuel Schmidt; Maciek R Antoniewicz; Mattheos A Koffas; Eleftherios T Papoutsakis

doi:10.1016/j.ymben.2016.10.015

Engineering the biological conversion of methanol to specialty chemicals in Escherichia coli

Metab Eng. 2017 Jan:39:49-59. doi: 10.1016/j.ymben.2016.10.015. Epub 2016 Nov 1.

Affiliations

¹ Department of Chemical and Biomolecular Engineering, University of Delaware, 150 Academy St., Newark, DE 19716, USA; The Delaware Biotechnology Institute, University of Delaware, 15 Innovation Way, Newark, DE 19711, USA. Electronic address: wbwhit@udel.edu.
² Department of Chemical and Biological Engineering, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, 110 8th St., Troy, NY 12180, USA; Department of Chemistry, Hamilton College, 198 College Hill Rd., Clinton, NY 13323, USA. Electronic address: jajones@hamilton.edu.
³ Department of Chemical and Biomolecular Engineering, University of Delaware, 150 Academy St., Newark, DE 19716, USA; The Delaware Biotechnology Institute, University of Delaware, 15 Innovation Way, Newark, DE 19711, USA. Electronic address: rkbenne@udel.edu.
⁴ Department of Chemical and Biomolecular Engineering, University of Delaware, 150 Academy St., Newark, DE 19716, USA. Electronic address: jgonz@udel.edu.
⁵ Department of Chemical and Biological Engineering, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, 110 8th St., Troy, NY 12180, USA. Electronic address: vernav@rpi.edu.
⁶ Department of Chemical and Biological Engineering, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, 110 8th St., Troy, NY 12180, USA. Electronic address: collis6@rpi.edu.
⁷ Department of Chemical and Biomolecular Engineering, University of Delaware, 150 Academy St., Newark, DE 19716, USA. Electronic address: mpalmer@udel.edu.
⁸ Department of Chemical and Biomolecular Engineering, University of Delaware, 150 Academy St., Newark, DE 19716, USA. Electronic address: schm@udel.edu.
⁹ Department of Chemical and Biomolecular Engineering, University of Delaware, 150 Academy St., Newark, DE 19716, USA. Electronic address: mranton@udel.edu.
¹⁰ Department of Chemical and Biological Engineering, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, 110 8th St., Troy, NY 12180, USA; Department of Biological Sciences, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, 110 8th St., Troy, NY 12180, USA. Electronic address: koffam@rpi.edu.
¹¹ Department of Chemical and Biomolecular Engineering, University of Delaware, 150 Academy St., Newark, DE 19716, USA; The Delaware Biotechnology Institute, University of Delaware, 15 Innovation Way, Newark, DE 19711, USA. Electronic address: papoutsakis@dbi.udel.edu.

PMID: 27815193
DOI: 10.1016/j.ymben.2016.10.015

Abstract

Methanol is an attractive substrate for biological production of chemicals and fuels. Engineering methylotrophic Escherichia coli as a platform organism for converting methanol to metabolites is desirable. Prior efforts to engineer methylotrophic E. coli were limited by methanol dehydrogenases (Mdhs) with unfavorable enzyme kinetics. We engineered E. coli to utilize methanol using a superior NAD-dependent Mdh from Bacillus stearothermophilus and ribulose monophosphate (RuMP) pathway enzymes from B. methanolicus. Using ¹³C-labeling, we demonstrate this E. coli strain converts methanol into biomass components. For example, the key TCA cycle intermediates, succinate and malate, exhibit labeling up to 39%, while the lower glycolytic intermediate, 3-phosphoglycerate, up to 53%. Multiple carbons are labeled for each compound, demonstrating a cycling RuMP pathway for methanol assimilation to support growth. By incorporating the pathway to synthesize the flavanone naringenin, we demonstrate the first example of in vivo conversion of methanol into a specialty chemical in E. coli.

Keywords: E. coli; Methanol; Methylotrophy; Pathway engineering for substrate utilization.

MeSH terms

Alcohol Oxidoreductases / genetics*
Alcohol Oxidoreductases / metabolism
Biosynthetic Pathways / physiology
Escherichia coli / physiology*
Escherichia coli Proteins / genetics
Escherichia coli Proteins / metabolism*
Flavanones / biosynthesis*
Flavanones / genetics
Genetic Enhancement / methods
Metabolic Engineering / methods*
Metabolic Networks and Pathways / physiology*
Methanol / metabolism*

Substances

Escherichia coli Proteins
Flavanones
Alcohol Oxidoreductases
alcohol dehydrogenase (acceptor)
naringenin
Methanol