title: NMR Binding and Functional Assays for Detecting Inhibitors of S. aureus MnaA.
abstract: Nonessential enzymes in the staphylococcal wall teichoic acid (WTA) pathway serve as highly validated β-lactam potentiation targets. MnaA (UDP-GlcNAc 2-epimerase) plays an important role in an early step of WTA biosynthesis by providing an activated form of ManNAc. Identification of a selective MnaA inhibitor would provide a tool to interrogate the contribution of the MnaA enzyme in the WTA pathway as well as serve as an adjuvant to restore β-lactam activity against methicillin-resistant Staphylococcus aureus (MRSA). However, development of an epimerase functional assay can be challenging since both MnaA substrate and product (UDP-GlcNAc/UDP-ManNAc) share an identical molecular weight. Herein, we developed a nuclear magnetic resonance (NMR) functional assay that can be combined with other NMR approaches to triage putative MnaA inhibitors from phenotypic cell-based screening campaigns. In addition, we determined that tunicamycin, a potent WTA pathway inhibitor, inhibits both S. aureus MnaA and a functionally redundant epimerase, Cap5P.
authors.affiliation: Structural Chemistry, Merck Research Laboratories, Kenilworth, NJ, USA.
authors.forename: Yan
authors.initials: Y
authors.lastname: Hou
authors.affiliation: Pharmacology, Biochem & Biophysics, Merck Research Laboratories, Kenilworth, NJ, USA.
authors.forename: Todd
authors.initials: T
authors.lastname: Mayhood
authors.affiliation: Pharmacology, Biochem & Biophysics, Merck Research Laboratories, Kenilworth, NJ, USA.
authors.forename: Payal
authors.initials: P
authors.lastname: Sheth
authors.affiliation: Discovery-Biology, Infectious Diseases, Merck Research Laboratories, Kenilworth, NJ, USA.
authors.forename: Christopher M
authors.initials: CM
authors.lastname: Tan
authors.affiliation: Exploratory Chemistry, Merck Research Laboratories, Kenilworth, NJ, USA.
authors.forename: Marc
authors.initials: M
authors.lastname: Labroli
authors.affiliation: Exploratory Chemistry, Merck Research Laboratories, Kenilworth, NJ, USA.
authors.forename: Jing
authors.initials: J
authors.lastname: Su
authors.affiliation: Structural Chemistry, Merck Research Laboratories, Kenilworth, NJ, USA.
authors.forename: Daniel F
authors.initials: DF
authors.lastname: Wyss
authors.affiliation: Discovery-Biology, Infectious Diseases, Merck Research Laboratories, Kenilworth, NJ, USA.
authors.forename: Terry
authors.initials: T
authors.lastname: Roemer
authors.affiliation: Structural Chemistry, Merck Research Laboratories, Kenilworth, NJ, USA mark.mccoy@merck.com.
authors.forename: Mark A
authors.initials: MA
authors.lastname: McCoy
mesh_terms: D002238:Carbohydrate Epimerases / Q000037:antagonists & inhibitors / Q000737:chemistry; D002473:Cell Wall / Q000737:chemistry / Q000187:drug effects*; D006801:Humans; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D055624:Methicillin-Resistant Staphylococcus aureus / Q000187:drug effects* / Q000472:pathogenicity; D013682:Teichoic Acids / Q000737:chemistry / Q000378:metabolism; D014539:Uridine Diphosphate Sugars / Q000737:chemistry / Q000378:metabolism; D018440:beta-Lactam Resistance / Q000187:drug effects; D001618:beta-Lactamases / Q000737:chemistry / Q000187:drug effects
chemical_list: D013682:Teichoic Acids; D014539:Uridine Diphosphate Sugars; C059191:uridine diphosphate N-acetylmannosamine; D001618:beta-Lactamases; D002238:Carbohydrate Epimerases; C100033:UDP acetylglucosamine-2-epimerase
keywords: MnaA (UDP-GlcNAc 2-epimerase); NMR-based hit validation; drug discovery; tunicamycin; wall teichoic acid (WTA)