title: A propionate-inducible expression system based on the Corynebacterium glutamicum prpD2 promoter and PrpR activator and its application for the redirection of amino acid biosynthesis pathways.
abstract: A novel expression system for Corynebacterium glutamicum, based on the transcriptional activator of propionate metabolism genes PrpR and its target promoter/operator sequence, was developed and tested. The activator PrpR is co-activated by propionate added to the growth medium. In a minimal medium a propionate concentration of only 1 mg l⁻¹ was found to be sufficient for full induction (up to 120-fold) of its native target, the propionate metabolism operon prpDBC2. Then, an artificial transcription and translation reporter system, using the cat gene encoding chloramphenicol acetyl transferase was constructed and tested. The induction was found to be as fast and as high as in the natural system, reaching its maximal transcriptional induction rate within 2 min and a significant accumulation of Cat protein at around 30 min. The duration of the induced transcription was found to be controllable by the propionate concentration applied. The prpD2 promoter and PrpR activator based expression system revealed very similar characteristics in minimal and complex media, making it ideal for applications in large scale industrial fermentations. As a proof-of-principle, the expression system was employed for the propionate-inducible redirection of metabolites in a lysine-production C. glutamicum strain at the homoserine dehydrogenase (hom) branching point, which resulted in an up to 2.5-fold increase of the concentrations of the three amino acids (threonine, homoserine and isoleucine) in the supernatant.
authors.affiliation: Technologieplattform Genomik, Centrum für Biotechnologie, Universität Bielefeld, Universitätsstr. 27, 33615 Bielefeld, Germany.
authors.forename: Jens K
authors.initials: JK
authors.lastname: Plassmeier
authors.forename: Tobias
authors.initials: T
authors.lastname: Busche
authors.forename: Stella
authors.initials: S
authors.lastname: Molck
authors.forename: Marcus
authors.initials: M
authors.lastname: Persicke
authors.forename: Alfred
authors.initials: A
authors.lastname: Pühler
authors.forename: Christian
authors.initials: C
authors.lastname: Rückert
authors.forename: Jörn
authors.initials: J
authors.lastname: Kalinowski
mesh_terms: D000596:Amino Acids / Q000096:biosynthesis* / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics; D057486:Carbon Cycle; D002851:Chromatography, High Pressure Liquid; D002951:Citrates / Q000378:metabolism; D048230:Corynebacterium glutamicum / Q000235:genetics* / Q000378:metabolism*; D005285:Fermentation; D008401:Gas Chromatography-Mass Spectrometry; D005798:Genes, Bacterial; D060847:Metabolic Engineering / Q000379:methods*; D053858:Metabolic Networks and Pathways; D009876:Operon; D011401:Promoter Regions, Genetic; D011422:Propionates / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics*
chemical_list: D000596:Amino Acids; D001426:Bacterial Proteins; D002951:Citrates; D011422:Propionates; D014157:Transcription Factors; C031605:2-methylcitric acid