Bifunctional nitrone-conjugated secondary metabolite targeting the ribosome

Brian O. Bachmann, Daniel N. Wilson, Emilianne M. Limbrick, Michael Graf, Dagmara K. Derewacz, Fabian Nguyen, Jeffrey M. Spraggins, Maximiliane Wieland, Audrey E. Ynigez-Gutierrez, Benjamin J. Reisman, Boris Zinshteyn, Kathryn M. McCulloch, T. M. Iverson, Rachel Green

Research output: Contribution to journalArticlepeer-review

1 Scopus citations


Many microorganisms possess the capacity for producing multiple antibiotic secondary metabolites. In a few notable cases, combinations of secondary metabolites produced by the same organism are used in important combination therapies for treatment of drug-resistant bacterial infections. However, examples of conjoined roles of bioactive metabolites produced by the same organism remain uncommon. During our genetic functional analysis of oxidase-encoding genes in the everninomicin producer Micromonospora carbonacea var. aurantiaca, we discovered previously uncharacterized antibiotics everninomicin N and O, comprised of an everninomicin fragment conjugated to the macrolide rosamicin via a rare nitrone moiety. These metabolites were determined to be hydrolysis products of everninomicin P, a nitrone-linked conjugate likely the result of nonenzymatic condensation of the rosamicin aldehyde and the octasaccharide everninomicin F, possessing a hydroxylamino sugar moiety. Rosamicin binds the erythromycin macrolide binding site approximately 60 Å from the orthosomycin binding site of everninomicins. However, while individual ribosomal binding sites for each functional half of everninomicin P are too distant for bidentate binding, ligand displacement studies demonstrated that everninomicin P competes with rosamicin for ribosomal binding. Chemical protection studies and structural analysis of everninomicin P revealed that everninomicin P occupies both the macrolide- and orthosomycin-binding sites on the 70S ribosome. Moreover, resistance mutations within each binding site were overcome by the inhibition of the opposite functional antibiotic moiety binding site. These data together demonstrate a strategy for coupling orthogonal antibiotic pharmacophores, a surprising tolerance for substantial covalent modification of each antibiotic, and a potential beneficial strategy to combat antibiotic resistance.

Original languageEnglish (US)
Pages (from-to)18369-18377
Number of pages9
JournalJournal of the American Chemical Society
Issue number43
StatePublished - Oct 28 2020

ASJC Scopus subject areas

  • Catalysis
  • Chemistry(all)
  • Biochemistry
  • Colloid and Surface Chemistry


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