Journal of Applied Biological Chemistry

The Korean Society for Applied Biological Chemistry (한국응용생명화학회)
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Cytochrome P450 and the glycosyltransferase genes are necessary for product release from epipyrone polyketide synthase in Epicoccum nigrum

  • Choi, Eun Ha (Department of Biological Sciences and Bioinformatics, Myongji University) ;
  • Park, Si-Hyung (Department of Oriental Medicine Resources and Institute for Traditional Korean Medicine Industry, Mokpo National University) ;
  • Kwon, Hyung-Jin (Department of Biological Sciences and Bioinformatics, Myongji University)
  • Received : 2021.06.17
  • Accepted : 2021.07.15
  • Published : 2021.09.30
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Abstract

The epipyrone (EPN) biosynthetic gene cluster of Epicoccum nigrum is composed of epnC, epnB, and epnA, which encode cytochrome P450 oxidase, glycosyltransferase, and highly reducing polyketide synthase, respectively. Gene inactivation mutants for epnA, epnB, and epnC were previously generated, and it was found that all of them were incapable of producing EPN and any of its related compounds. It was also reported that epnB inactivation abolished epnA transcription, generating ΔepnAB. This study shows that the introduction of native epnC readily restored EPN production in ΔepnC, suggesting that epnC is essential for polyketide release from EpnA and implies that EpnC works during the polyketide chain assembly of EpnA. Introduction of epnC promoter-epnA restored EPN production in ΔepnA. The ΔepnB genotype was prepared by introducing the epnA expression vector into ΔepnAB, and it was found that the resulting recombinant strain did not produce any EPN-related compounds. A canonical epnB inactivation strain was also generated by deleting its 5'-end. At the deletion point, an Aspergllus nidulans gpdA promoter was inserted to ensure the transcription of epnA, which is located downstream of epnB. Examination of the metabolite profile of the resulting ΔepnB mutant via LC-mass spectrometry verified that no EPN-related compound was produced in this strain. This substantiates that C-glycosylation by EpnB is a prerequisite for the release of EpnA-tethered product. In conclusion, it is proposed that cytochrome P450 oxidase and glycosyltransferase work in concert with polyketide synthase to generate EPN without the occurrence of any free intermediates.

Keywords

Acknowledgement

This research was supported by Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education (NRF-2016R1D1A1B02009237).

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