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Characterisation of the genes involved in the biosynthesis and attachment of the aminodeoxysugar d-forosamine in the auricin gene cluster of Streptomyces aureofaciens CCM3239

  • Applied genetics and molecular biotechnology
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Abstract

We previously identified the aur1 gene cluster which produces the angucycline antibiotic auricin. Preliminary characterisation of auricin revealed that it is modified by a single aminodeoxysugar, d-forosamine. Here we characterise the d-forosamine-specific genes. The four close tandem genes, aur1TQSV, encoding enzymes involved in the initial steps of the deoxysugar biosynthesis, were located on a large operon with other core auricin biosynthetic genes. Deleting these genes resulted in the absence of auricin and the production of deglycosylated auricin intermediates. The two final d-forosamine biosynthetic genes, sa59, an NDP-hexose aminotransferase, and sa52, an NDP-aminohexose N-dimethyltransferase, are located in a region rather distant from the core auricin genes. A deletion analysis of these genes confirmed their role in d-forosamine biosynthesis. The Δsa59 mutant had a phenotype similar to that of the cluster deletion mutant, while the Δsa52 mutant produced an auricin with a demethylated d-forosamine. Although auricin contains a single deoxyhexose, two glycosyltransferase genes were found to participate in the attachment of d-forosamine to the auricin aglycon. An analysis of the expression of the d-forosamine biosynthesis genes revealed that the initial d-forosamine biosynthetic genes aur1TQSV are regulated together with the other auricin core genes by the aur1Ap promoter under the control of the auricin-specific activator Aur1P. The expression of the other d-forosamine genes, however, is governed by promoters differentially dependent upon the two SARP family auricin-specific activators Aur1PR3 and Aur1PR4. These promoters contain direct repeats similar to the SARP consensus sequence and are involved in the interaction with both regulators.

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Acknowledgments

We are grateful to Dr. Bertold Gust (John Innes Centre, Norwich, UK) for kindly providing all the plasmids and strains used in the PCR targeting system; the system itself was supplied by Plant Bioscience Ltd. (Norwich, UK). This work was supported by the Slovak Research and Development Agency under contract No. APVV-0203-11. The research leading to these results received funding from the European Commission’s Seventh Framework Programme (FP7/2007-2013) under the grant agreement STREPSYNTH (project No. 613877). This work was co-funded by the Slovak Research and Development Agency under contract No. DO7RP-0037-12.

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  1. Institute of Molecular Biology, Slovak Academy of Sciences, Dubravska cesta 21, 845 51, Bratislava, Slovak Republic

    Carmen Bekeova, Alena Rehakova, Lubomira Feckova, Renata Novakova, Erik Mingyar & Jan Kormanec

  2. Institute of Chemistry, Slovak Academy of Sciences, 845 38, Bratislava, Slovak Republic

    Silvia Vlckova

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  1. Carmen Bekeova
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  2. Alena Rehakova
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Correspondence to Jan Kormanec.

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Bekeova, C., Rehakova, A., Feckova, L. et al. Characterisation of the genes involved in the biosynthesis and attachment of the aminodeoxysugar d-forosamine in the auricin gene cluster of Streptomyces aureofaciens CCM3239. Appl Microbiol Biotechnol 100, 3177–3195 (2016). https://doi.org/10.1007/s00253-015-7214-9

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