Leveraging host metabolism for bisdemethoxycurcumin production in Pseudomonas putida

Matthew R. Incha; Mitchell G. Thompson; Jacquelyn M. Blake-Hedges; Yuzhong Liu; Allison N. Pearson; Matthias Schmidt; Jennifer W. Gin; Christopher J. Petzold; Adam M. Deutschbauer; Jay D. Keasling

Metabolic Engineering Communications (Jun 2020)

Leveraging host metabolism for bisdemethoxycurcumin production in Pseudomonas putida

Matthew R. Incha,
Mitchell G. Thompson,
Jacquelyn M. Blake-Hedges,
Yuzhong Liu,
Allison N. Pearson,
Matthias Schmidt,
Jennifer W. Gin,
Christopher J. Petzold,
Adam M. Deutschbauer,
Jay D. Keasling

Affiliations

Matthew R. Incha: Joint BioEnergy Institute, 5885 Hollis Street, Emeryville, CA, 94608, USA; Biological Systems & Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, CA, 94720, USA; Department of Plant and Microbial Biology, University of California, Berkeley, CA, 94720, USA
Mitchell G. Thompson: Joint BioEnergy Institute, 5885 Hollis Street, Emeryville, CA, 94608, USA; Biological Systems & Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, CA, 94720, USA; Department of Plant and Microbial Biology, University of California, Berkeley, CA, 94720, USA
Jacquelyn M. Blake-Hedges: Joint BioEnergy Institute, 5885 Hollis Street, Emeryville, CA, 94608, USA; Biological Systems & Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, CA, 94720, USA; Department of Chemistry, University of California, Berkeley, CA, 94720, USA
Yuzhong Liu: Joint BioEnergy Institute, 5885 Hollis Street, Emeryville, CA, 94608, USA; Biological Systems & Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, CA, 94720, USA
Allison N. Pearson: Joint BioEnergy Institute, 5885 Hollis Street, Emeryville, CA, 94608, USA; Biological Systems & Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, CA, 94720, USA
Matthias Schmidt: Joint BioEnergy Institute, 5885 Hollis Street, Emeryville, CA, 94608, USA; Biological Systems & Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, CA, 94720, USA
Jennifer W. Gin: Joint BioEnergy Institute, 5885 Hollis Street, Emeryville, CA, 94608, USA; Biological Systems & Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, CA, 94720, USA
Christopher J. Petzold: Joint BioEnergy Institute, 5885 Hollis Street, Emeryville, CA, 94608, USA; Biological Systems & Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, CA, 94720, USA
Adam M. Deutschbauer: Department of Plant and Microbial Biology, University of California, Berkeley, CA, 94720, USA; Environmental Genomics and Systems Biology Division, Lawrence Berkeley National Laboratory, Berkeley, CA, USA
Jay D. Keasling: Joint BioEnergy Institute, 5885 Hollis Street, Emeryville, CA, 94608, USA; Biological Systems & Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, CA, 94720, USA; Department of Chemical and Biomolecular Engineering, University of California, Berkeley, CA, 94720, USA; The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Denmark; Center for Synthetic Biochemistry, Institute for Synthetic Biology, Shenzhen Institutes for Advanced Technologies, Shenzhen, China; Corresponding author. Department of Chemical and Biomolecular Engineering, University of California, Berkeley, CA, 94720, USA.

Journal volume & issue: Vol. 10
p. e00119

Abstract

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Pseudomonas putida is a saprophytic bacterium with robust metabolisms and strong solvent tolerance making it an attractive host for metabolic engineering and bioremediation. Due to its diverse carbon metabolisms, its genome encodes an array of proteins and enzymes that can be readily applied to produce valuable products. In this work we sought to identify design principles and bottlenecks in the production of type III polyketide synthase (T3PKS)-derived compounds in P. putida. T3PKS products are widely used as nutraceuticals and medicines and often require aromatic starter units, such as coumaroyl-CoA, which is also an intermediate in the native coumarate catabolic pathway of P. putida. Using a randomly barcoded transposon mutant (RB-TnSeq) library, we assayed gene functions for a large portion of aromatic catabolism, confirmed known pathways, and proposed new annotations for two aromatic transporters. The 1,3,6,8-tetrahydroxynapthalene synthase of Streptomyces coelicolor (RppA), a microbial T3PKS, was then used to rapidly assay growth conditions for increased T3PKS product accumulation. The feruloyl/coumaroyl CoA synthetase (Fcs) of P. putida was used to supply coumaroyl-CoA for the curcuminoid synthase (CUS) of Oryza sativa, a plant T3PKS. We identified that accumulation of coumaroyl-CoA in this pathway results in extended growth lag times in P. putida. Deletion of the second step in coumarate catabolism, the enoyl-CoA hydratase-lyase (Ech), resulted in increased production of the type III polyketide bisdemethoxycurcumin.

Published in Metabolic Engineering Communications

ISSN: 2214-0301 (Online)
Publisher: Elsevier
Country of publisher: Netherlands
LCC subjects: Technology: Chemical technology: Biotechnology; Science: Biology (General)
Website: http://www.journals.elsevier.com/metabolic-engineering-communications/

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