3-Hydroxylation of the polycyclic tetramate macrolactam in the biosynthesis of antifungal HSAF from Lysobacter enzymogenes C3

Yaoyao Li, Justin Huffman, Yu Li, Liangcheng Du, Yuemao Shen

Research output: Contribution to journalArticle

29 Scopus citations

Abstract

HSAF isolated from Lysobacter enzymogenes is an antifungal natural product with a new mode of action. The polycyclic tetramate macrolactam (PTM) carries a 3-hydroxyl group on the ornithine residue, which is a feature common to nearly all PTM type natural products found in phylogenetically diverse organisms. A previous gene disruption experiment indicated that the sterol desaturase (SD) gene, which is clustered with the central polyketide synthase-nonribosomal peptide synthetase (PKS-NRPS), was involved in the 3-hydroxylation. However, the mechanism for this hydroxylation had not been established. Here, we determined the structure of the main metabolite accumulated in the SD mutant, which is 3-dehydroxy HSAF (3-deOH-HSAF). This compound lost the antifungal activity against Penicillium avellaneum, showing the crucial role of the 3-hydroxyl group. We then expressed the SD gene in E. coli. Upon feeding 3-deOH-HSAF, E. coli produced HSAF. When the SD enzyme extract was incubated with 3-deOH-HSAF in the presence of NADPH, HSAF was also produced. The results demonstrated that the SD gene encodes a 3-hydroxylase of the HSAF carbon chain. In addition, the data support that the 3-hydroxylation step is likely a post-PKS-NRPS event in the HSAF biosynthetic pathway. Finally, we co-expressed the ferredoxin reductase (FNR) gene, which is also clustered with the PKS-NRPS gene, with the SD gene in E. coli. The results showed that FNR significantly enhanced the conversion of 3-deOH-HSAF into HSAF. Together, the study established the mechanism for the installation of this common functionality that is important to the activity of the PTM-type natural products.

Original languageEnglish (US)
Pages (from-to)982-986
Number of pages5
JournalMedChemComm
Volume3
Issue number8
DOIs
StatePublished - Aug 2012

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Medicine
  • Pharmacology
  • Pharmaceutical Science
  • Drug Discovery
  • Organic Chemistry

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