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Methyltransferase Contingencies in the Pathway of Everninomicin D Antibiotics and Analogues


Limbrick EM , Yñigez-Gutierrez AE , Dulin CC , Derewacz DK , Spraggins JM , McCulloch KM , Iverson TM , Bachmann BO , . Chembiochem : a European journal of chemical biology. 2020 9 7; 21(23). 3349-3358


Everninomicins are orthoester oligosaccharide antibiotics with potent activity against multidrug-resistant bacterial pathogens. Everninomicins act by disrupting ribosomal assembly in a distinct region in comparison to clinically prescribed drugs. We employed microporous intergeneric conjugation with Escherichia coli to manipulate Micromonospora for targeted gene-replacement studies of multiple putative methyltransferases across the octasaccharide scaffold of everninomicin effecting the A , C, F, and H rings. Analyses of gene-replacement and genetic complementation mutants established the mutability of the everninomicin scaffold through the generation of 12 previously unreported analogues and, together with previous results, permitted assignment of the ten methyltransferases required for everninomicin biosynthesis. The in vitro activity of A – and H-ring-modifying methyltransferases demonstrated the ability to catalyze late-stage modification of the scaffold on an A -ring phenol and H-ring C-4′ hydroxy moiety. Together these results establish the potential of the everninomicin scaffold for modification through mutagenesis and in vitro modification of advanced biosynthetic intermediates.