title: A novel reporter system for bacterial and mammalian cells based on the non-ribosomal peptide indigoidine.
abstract: The biosynthesis of non-ribosomal peptides, many of which have pharmaceutical activities, is an evolutionary privilege of microorganisms. Capitalizing on the universal set of the Streptomyces lavendulae non-ribosomal peptide synthase BpsA and the Streptomyces verticillus 4'-phosphopantetheinyl transferase Svp, we have engineered Escherichia coli as well as mammalian cells, including human stem cells, to produce the blue 3,3'-bipyridyl pigment keto-indigoidine and the reduced colorless but fluorescent leuco-isoform. Detailed characterization of a tailored substrate-free chromogenic assay and FACS analysis showed that indigoidine's blue color and fluorescence could be reliably profiled in bacteria and mammalian cells using standard multiwell-compatible detection equipment. Besides serving as an inexpensive, reliable, versatile and easy-to-assay cross-kingdom reporter system, the potential of having mammalian cells produce non-ribosomal peptides, preferably ones with biopharmaceutical activities, may provide novel treatment opportunities in future gene- and cell-based therapies.
authors.affiliation: Department of Biosystems Science and Engineering, ETH Zürich, Mattenstrasse 26, CH-4058 Basel, Switzerland.
authors.forename: Marius
authors.initials: M
authors.lastname: Müller
authors.forename: Simon
authors.initials: S
authors.lastname: Ausländer
authors.forename: David
authors.initials: D
authors.lastname: Ausländer
authors.forename: Christian
authors.initials: C
authors.lastname: Kemmer
authors.forename: Martin
authors.initials: M
authors.lastname: Fussenegger
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D053595:Embryonic Stem Cells; D004926:Escherichia coli / Q000378:metabolism; D017930:Genes, Reporter; D006801:Humans; D060847:Metabolic Engineering / Q000379:methods; D010453:Peptide Synthases / Q000235:genetics / Q000378:metabolism; D010881:Piperidones / Q000378:metabolism*; D013302:Streptomyces / Q000201:enzymology*; D017855:Transferases (Other Substituted Phosphate Groups) / Q000235:genetics / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D010881:Piperidones; C082662:phosphopantetheinyl transferase; C446045:indigoidine; D017855:Transferases (Other Substituted Phosphate Groups); D010453:Peptide Synthases; C104385:non-ribosomal peptide synthase