High titer methyl ketone production with tailored Pseudomonas taiwanensis VLB120

Salome C Nies; Tobias B Alter; Sophia Nölting; Susanne Thiery; An N T Phan; Noud Drummen; Jay D Keasling; Lars M Blank; Birgitta E Ebert

doi:10.1016/j.ymben.2020.08.003

High titer methyl ketone production with tailored Pseudomonas taiwanensis VLB120

Metab Eng. 2020 Nov:62:84-94. doi: 10.1016/j.ymben.2020.08.003. Epub 2020 Aug 15.

Authors

Salome C Nies¹, Tobias B Alter¹, Sophia Nölting¹, Susanne Thiery¹, An N T Phan¹, Noud Drummen¹, Jay D Keasling², Lars M Blank¹, Birgitta E Ebert³

Affiliations

¹ iAMB-Institute of Applied Microbiology, ABBt-Aachen Biology and Biotechnology, RWTH Aachen University, DE, Germany.
² Joint BioEnergy Institute, Emeryville, CA, 94608, USA; Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, 2800, Lyngby, Denmark; Lawrence Berkeley National Laboratory, Biological Systems and Engineering Division, Berkeley, CA, 94720, USA; Virtual Institute of Microbial Stress and Survival, Lawrence Berkeley National Laboratory, Berkeley, CA, 94720, USA; Physical Biosciences Division, Lawrence Berkeley National Laboratory, Berkeley, CA, 94720, USA; Dept. of Bioengineering, University of California, Berkeley, CA, 94720, USA; Dept. of Chemical Engineering, University of California, Berkeley, CA, 94720, USA; Synthetic Biochemistry Center, Institute for Synthetic Biology, Shenzhen Institutes for Advanced Technologies, Shenzhen, China.
³ iAMB-Institute of Applied Microbiology, ABBt-Aachen Biology and Biotechnology, RWTH Aachen University, DE, Germany; Australian Institute for Bioengineering and Nanotechnology (AIBN), The University of Queensland, Brisbane, QLD, 4072, Australia; CSIRO Future Science Platform in Synthetic Biology, Commonwealth Scientific and Industrial Research Organisation (CSIRO), Black Mountain, ACT, 2601, Australia. Electronic address: birgitta.ebert@uq.edu.au.

PMID: 32810591
DOI: 10.1016/j.ymben.2020.08.003

Abstract

Methyl ketones present a group of highly reduced platform chemicals industrially produced from petroleum-derived hydrocarbons. They find applications in the fragrance, flavor, pharmacological, and agrochemical industries, and are further discussed as biodiesel blends. In recent years, intense research has been carried out to achieve sustainable production of these molecules by re-arranging the fatty acid metabolism of various microbes. One challenge in the development of a highly productive microbe is the high demand for reducing power. Here, we engineered Pseudomonas taiwanensis VLB120 for methyl ketone production as this microbe has been shown to sustain exceptionally high NAD(P)H regeneration rates. The implementation of published strategies resulted in 2.1 g L_aq^-1 methyl ketones in fed-batch fermentation. We further increased the production by eliminating competing reactions suggested by metabolic analyses. These efforts resulted in the production of 9.8 g L_aq^-1 methyl ketones (corresponding to 69.3 g L_org^-1 in the in situ extraction phase) at 53% of the maximum theoretical yield. This represents a 4-fold improvement in product titer compared to the initial production strain and the highest titer of recombinantly produced methyl ketones reported to date. Accordingly, this study underlines the high potential of P. taiwanensis VLB120 to produce methyl ketones and emphasizes model-driven metabolic engineering to rationalize and accelerate strain optimization efforts.

Keywords: Biodiesel; Metabolic engineering; Metabolic modeling; Pseudomonads; Synthetic biology; Thioesterase.

Publication types

Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

Acetone
Fermentation
Metabolic Engineering*
Pseudomonas* / genetics

Substances

Acetone

Supplementary concepts

Pseudomonas taiwanensis