Synthetic Promoter Library for Modulation of Actinorhodin Production in Streptomyces coelicolor A3(2)

Sujata Vijay Sohoni, Alessandro Fazio, Christopher Workman, Ivan Mijakovic, Anna Eliasson Lantz

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    Abstract

    The objective of this study was the application of the synthetic promoter library (SPL) technology for modulation of actinorhodin production in Streptomyces coelicolor A3(2). The SPL technology was used to optimize the expression of a pathway specific positive transcriptional regulator Actll orf4, which activates the transcription of the S. coelicolor actinorhodin biosynthetic gene cluster. The native actll orf4 promoter was replaced with synthetic promoters, generating a S. coelicolor library with a broad range of expression levels of actll orf4. The resulting library was screened based on the yield of actinorhodin. Selected strains were further physiologically characterized. One of the strains from the library, ScoSPL20, showed considerably higher yield of actinorhodin and final actinorhodin titer, compared to S. coelicolor wild type and S. coelicolor with actll orf4 expressed from a strong constitutive promoter. ScoSPL20 demonstrated exceptional productivity despite having a comparatively weak expression from the promoter. Interestingly, the ScoSPL20 promoter was activated at a much earlier stage of growth compared to the wild type, demonstrating the advantage of fine-tuning and temporal tuning of gene expression in metabolic engineering. Transcriptome studies were performed in exponential and actinorhodin-producing phase of growth to compare gene expression between ScoSPL20 and the wild type. To our knowledge, this is the first successful application of the SPL technology for secondary metabolite production in filamentous bacteria.
    Original languageEnglish
    Article numbere99701
    JournalPLOS ONE
    Volume9
    Issue number6
    Number of pages12
    ISSN1932-6203
    DOIs
    Publication statusPublished - 2014

    Bibliographical note

    This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

    Keywords

    • Actinomycetes and Related Organisms Eubacteria Bacteria Microorganisms (Bacteria, Eubacteria, Microorganisms) - Streptomycetes and Related Genera [08840] Streptomyces coelicolor species strain-A3(2)
    • Facultatively Anaerobic Gram-Negative Rods Eubacteria Bacteria Microorganisms (Bacteria, Eubacteria, Microorganisms) - Enterobacteriaceae [06702] Escherichia coli species strain-DH5alpha
    • Streptomyces coelicolor actII orf4 gene [Streptomycetes and Related Genera] promoter, expression
    • Streptomyces coelicolor actinorhodin gene cluster [Streptomycetes and Related Genera] expression
    • Streptomyces coelicolor ScoSPL20 gene [Streptomycetes and Related Genera] promoter, expression
    • ActII
    • actinorhodin 1397-77-9
    • orf4
    • synthetic promoter library
    • 03502, Genetics - General
    • 31000, Physiology and biochemistry of bacteria
    • 31500, Genetics of bacteria and viruses
    • 39008, Food microbiology - General and miscellaneous
    • Biochemistry and Molecular Biophysics
    • Bioprocess Engineering
    • Molecular Genetics
    • MULTIDISCIPLINARY
    • ANTIBIOTIC PRODUCTION
    • SECONDARY METABOLISM
    • TRANSCRIPTIONAL ACTIVATION
    • BIOSYNTHETIC GENES
    • LIVIDANS
    • REGULATOR
    • EXPRESSION
    • SYSTEM
    • STRESS
    • RESISTANCE

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