title: Biosensor-driven adaptive laboratory evolution of l-valine production in Corynebacterium glutamicum.
abstract: Adaptive laboratory evolution has proven a valuable strategy for metabolic engineering. Here, we established an experimental evolution approach for improving microbial metabolite production by imposing an artificial selective pressure on the fluorescent output of a biosensor using fluorescence-activated cell sorting. Cells showing the highest fluorescent output were iteratively isolated and (re-)cultivated. The L-valine producer Corynebacterium glutamicum ΔaceE was equipped with an L-valine-responsive sensor based on the transcriptional regulator Lrp of C. glutamicum. Evolved strains featured a significantly higher growth rate, increased L-valine titers (~25%) and a 3-4-fold reduction of by-product formation. Genome sequencing resulted in the identification of a loss-of-function mutation (UreD-E188*) in the gene ureD (urease accessory protein), which was shown to increase L-valine production by up to 100%. Furthermore, decreased L-alanine formation was attributed to a mutation in the global regulator GlxR. These results emphasize biosensor-driven evolution as a straightforward approach to improve growth and productivity of microbial production strains.
authors.affiliation: IBG-1: Biotechnology, Forschungszentrum Jülich GmbH, 52425 Jülich, Germany.
authors.forename: Regina
authors.initials: R
authors.lastname: Mahr
authors.affiliation: IBG-1: Biotechnology, Forschungszentrum Jülich GmbH, 52425 Jülich, Germany.
authors.forename: Cornelia
authors.initials: C
authors.lastname: Gätgens
authors.affiliation: IBG-1: Biotechnology, Forschungszentrum Jülich GmbH, 52425 Jülich, Germany.
authors.forename: Jochem
authors.initials: J
authors.lastname: Gätgens
authors.affiliation: IBG-1: Biotechnology, Forschungszentrum Jülich GmbH, 52425 Jülich, Germany.
authors.forename: Tino
authors.initials: T
authors.lastname: Polen
authors.affiliation: Center for Biotechnology (CeBiTec), Bielefeld University, 33615 Bielefeld, Germany.
authors.forename: Jörn
authors.initials: J
authors.lastname: Kalinowski
authors.affiliation: IBG-1: Biotechnology, Forschungszentrum Jülich GmbH, 52425 Jülich, Germany. Electronic address: j.frunzke@fz-juelich.de.
authors.forename: Julia
authors.initials: J
authors.lastname: Frunzke
mesh_terms: D015374:Biosensing Techniques; D003352:Corynebacterium / Q000235:genetics / Q000378:metabolism; D048230:Corynebacterium glutamicum / Q000235:genetics / Q000378:metabolism*; D004269:DNA, Bacterial / Q000235:genetics; D004274:DNA, Recombinant; D019020:Directed Molecular Evolution; D005456:Fluorescent Dyes; D060847:Metabolic Engineering; D009154:Mutation; D014633:Valine / Q000096:biosynthesis*
chemical_list: D004269:DNA, Bacterial; D004274:DNA, Recombinant; D005456:Fluorescent Dyes; D014633:Valine
keywords: Adaptive laboratory evolution; Biosensor; Corynebacterium glutamicum; L-valine; Metabolic engineering; Transcription factor