title: Combinatorial pathway engineering for optimized production of the anti-malarial FR900098.
abstract: As resistance to current anti-malarial therapeutics spreads, new compounds to treat malaria are increasingly needed. One promising compound is FR900098, a naturally occurring phosphonate. Due to limitations in both chemical synthesis and biosynthetic methods for FR900098 production, this potential therapeutic has yet to see widespread implementation. Here we applied a combinatorial pathway engineering strategy to improve the production of FR900098 in Escherichia coli by modulating each of the pathway's nine genes with four promoters of different strengths. Due to the large size of the library and the low screening throughput, it was necessary to develop a novel screening strategy that significantly reduced the sample size needed to find an optimal strain. This was done by using biased libraries that localize searching around top hits and home in on high-producing strains. By incorporating this strategy, a significantly improved strain was found after screening less than 3% of the entire library. When coupled with culturing optimization, a strain was found to produce 96 mg/L, a 16-fold improvement over the original strain. We believe the enriched library method developed here can be used on other large pathways that may be difficult to engineer by combinatorial methods due to low screening throughput.
authors.affiliation: Department of Chemical and Biomolecular Engineering and Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois.
authors.forename: Todd S
authors.initials: TS
authors.lastname: Freestone
authors.affiliation: Department of Chemical and Biomolecular Engineering and Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois. zhao5@illinois.edu.|Departments of Chemistry, Biochemistry, and Bioengineering, Center for Biophysics and Computational Biology, School of Molecular and Cellular Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois, 61801. zhao5@illinois.edu.
authors.forename: Huimin
authors.initials: H
authors.lastname: Zhao
mesh_terms: D000962:Antimalarials / Q000378:metabolism*; D001709:Biotechnology / Q000379:methods; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism*; D005820:Genetic Testing; D007218:Industrial Microbiology / Q000379:methods; D060847:Metabolic Engineering / Q000379:methods*; D053858:Metabolic Networks and Pathways / Q000235:genetics*; D063065:Organophosphonates / Q000378:metabolism*; D013678:Technology, Pharmaceutical / Q000379:methods*
chemical_list: D000962:Antimalarials; D063065:Organophosphonates
keywords: FR900098; anti-malarial; metabolic engineering; pathway engineering