title: Reconstituted biosynthesis of the nonribosomal macrolactone antibiotic valinomycin in Escherichia coli.
abstract: The structural complexity of nonribosomal peptides (NRPs) impeding economic chemical synthesis and poor cultivability of source organisms limits the development of bioprocesses for novel bioactive compounds. Since nonribosomal peptide synthetases (NRPSs) assemble NRPs from simple amino acid building blocks, heterologous expression of NRPSs in a robust and easy to manipulate expression host is an attractive strategy to make pharmaceutically relevant NRPs more accessible and is also a basis for engineering of these enzymes to generate novel synthetic bioactive compounds. Here we show a systematic approach for the heterologous expression of the 654 kDa heterodimeric valinomycin synthetase (VlmSyn) from Streptomyces tsusimaensis in a soluble and active form in Escherichia coli. VlmSyn activity and precursor requirements were determined in vitro and provided evidence for a previously proposed model of valinomycin biosynthesis. In vivo production of recombinant valinomycin, a macrolactone antibiotic with reported antifungal, antibacterial, and antiviral activities, was achieved using an engineered E. coli strain growing in inexpensive media and independent of the supplementation with precursors and further optimization of the cultivation conditions. Tailoring of VlmSyn in E. coli paves the way to the production of novel valinomycin analogues in the future.
authors.affiliation: Institut für Biotechnologie and ‡Institut für Chemie, Technische Universität Berlin , D-10623 Berlin, Germany.
authors.forename: Jennifer
authors.initials: J
authors.lastname: Jaitzig
authors.forename: Jian
authors.initials: J
authors.lastname: Li
authors.forename: Roderich D
authors.initials: RD
authors.lastname: Süssmuth
authors.forename: Peter
authors.initials: P
authors.lastname: Neubauer
mesh_terms: D000900:Anti-Bacterial Agents / Q000032:analysis / Q000096:biosynthesis* / Q000737:chemistry; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D013058:Mass Spectrometry; D010453:Peptide Synthases / Q000235:genetics / Q000378:metabolism; D013302:Streptomyces / Q000201:enzymology / Q000235:genetics; D014634:Valinomycin / Q000032:analysis / Q000096:biosynthesis* / Q000737:chemistry
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D014634:Valinomycin; D010453:Peptide Synthases; C104385:non-ribosomal peptide synthase