title: Enhancing poly(3-hydroxyalkanoate) production in Escherichia coli by the removal of the regulatory gene arcA.
abstract: Recombinant Escherichia coli is a desirable platform for the production of many biological compounds including poly(3-hydroxyalkanoates), a class of naturally occurring biodegradable polyesters with promising biomedical and material applications. Although the controlled production of desirable polymers is possible with the utilization of fatty acid feedstocks, a central challenge to this biosynthetic route is the improvement of the relatively low polymer yield, a necessary factor of decreasing the production costs. In this study we sought to address this challenge by deleting arcA and ompR, two global regulators with the capacity to inhibit the uptake and activation of exogenous fatty acids. We found that polymer yields in a ΔarcA mutant increased significantly with respect to the parental strain. In the parental strain, PHV yields were very low but improved 64-fold in the ΔarcA mutant (1.92-124 mg L-1) The ΔarcA mutant also allowed for modest increases in some medium chain length polymer yields, while weight average molecular weights improved by approximately 1.5-fold to 12-fold depending on the fatty acid substrate utilized. These results were supported by an analysis of differential gene expression, which showed that the key genes (fadD, fadL, and fadE) encoding fatty acid degradation enzymes were all upregulated by 2-, 10-, and 31-fold in an ΔarcA mutant, respectively. Additionally, the short chain length fatty acid uptake genes atoA, atoE and atoD were upregulated by 103-, 119-, and 303-fold respectively, though these values are somewhat inflated due to low expression in the parental strain. Overall, this study demonstrates that arcA is an important target to improve PHA production from fatty acids.
authors.affiliation: Department of Chemistry, State University of New York College of Environmental Science and Forestry, 1 Forestry Drive, Syracuse, NY, 13210, USA.
authors.forename: Ryan A
authors.initials: RA
authors.lastname: Scheel
authors.affiliation: Department of Chemistry, State University of New York College of Environmental Science and Forestry, 1 Forestry Drive, Syracuse, NY, 13210, USA.
authors.forename: Liyuan
authors.initials: L
authors.lastname: Ji
authors.affiliation: Department of Chemistry, State University of New York College of Environmental Science and Forestry, 1 Forestry Drive, Syracuse, NY, 13210, USA.
authors.forename: Benjamin R
authors.initials: BR
authors.lastname: Lundgren
authors.affiliation: Department of Chemistry, State University of New York College of Environmental Science and Forestry, 1 Forestry Drive, Syracuse, NY, 13210, USA. ctnomura@esf.edu.|Center for Applied Microbiology, State University of New York College of Environmental Science and Forestry, 1 Forestry Drive, Syracuse, NY, 13210, USA. ctnomura@esf.edu.|Hubei Collaborative Center for Green Transformation of Bio-Resources, College of Life Sciences, Hubei University, Wuhan, 430062, China. ctnomura@esf.edu.
authors.forename: Christopher T
authors.initials: CT
authors.lastname: Nomura
keywords: Beta-oxidation; Biodegradable polymer; Escherichia coli; Fatty acid metabolism; Polyhydroxyalkanoates; arcA