Use of an inducible promoter for antibiotic production in a heterologous host

Volker Dangel, Lucia Westrich, Margaret C. Smith, Lutz Heide, Bertolt Gust

Research output: Contribution to journalArticle

25 Citations (Scopus)

Abstract

The biosynthetic gene cluster of the aminocoumarin antibiotic novobiocin comprises 20 coding sequences. Sixteen of them code for essential enzymes for novobiocin formation, transcribed in the form of a single 18-kb polycistronic mRNA. In the present study, we replaced the genuine promoter of this operon by the tetracycline-inducible promoter tcp830 and at the same time deleting the two pathway-specific positive regulator genes of novobiocin biosynthesis. The heterologous producer Streptomyces coelicolor M512 harboring the modified gene cluster produced, upon addition of 2 mg L-1 of the inducer compound anhydrotetracyline, 3.4-fold more novobiocin than strains carrying the unmodified cluster. A second tcp830 promoter was inserted in the middle of the 18-kb operon in order to ensure adequate transcription of the rearmost genes. However, this did not lead to a further increase of novobiocin formation, showing that a single tcp830 promoter was sufficient to achieve high transcription of all 16 genes of the operon. A high induction of novobiocin formation was achieved within a wide range of anhydrotetracyline concentrations (0.25–2.0 mg L-1). Growth of the strains was not affected by these concentrations. The inducer compound could be added either at the time of inoculation or at any other time up to mid-growth phase, always achieving a similar final antibiotic production. Therefore, the tcp830 promoter presents a robust, easy-to-use system for the inducible expression of biosynthetic gene clusters in heterologous hosts, independent from the genuine regulatory network.
Original languageEnglish
Pages (from-to)261-269
Number of pages9
JournalApplied Microbiology and Biotechnology
Volume87
Issue number1
Early online date2 Feb 2010
DOIs
Publication statusPublished - Jun 2010

Fingerprint

Novobiocin
Anti-Bacterial Agents
Operon
Multigene Family
Aminocoumarins
Streptomyces coelicolor
Regulator Genes
Growth
Tetracycline
Genes
Messenger RNA
Enzymes

Keywords

  • novobiocin
  • inducible promoter
  • genetic engineering
  • streptomyces

Cite this

Use of an inducible promoter for antibiotic production in a heterologous host. / Dangel, Volker; Westrich, Lucia; Smith, Margaret C.; Heide, Lutz; Gust, Bertolt.

In: Applied Microbiology and Biotechnology, Vol. 87, No. 1, 06.2010, p. 261-269.

Research output: Contribution to journalArticle

Dangel, Volker ; Westrich, Lucia ; Smith, Margaret C. ; Heide, Lutz ; Gust, Bertolt. / Use of an inducible promoter for antibiotic production in a heterologous host. In: Applied Microbiology and Biotechnology. 2010 ; Vol. 87, No. 1. pp. 261-269.
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