title: Elucidation of DnaE as the Antibacterial Target of the Natural Product, Nargenicin.
abstract: Resistance to existing classes of antibiotics drives the need for discovery of novel compounds with unique mechanisms of action. Nargenicin A1, a natural product with limited antibacterial spectrum, was rediscovered in a whole-cell antisense assay. Macromolecular labeling in both Staphylococcus aureus and an Escherichia coli tolC efflux mutant revealed selective inhibition of DNA replication not due to gyrase or topoisomerase IV inhibition. S. aureus nargenicin-resistant mutants were selected at a frequency of ∼1 × 10(-9), and whole-genome resequencing found a single base-pair change in the dnaE gene, a homolog of the E. coli holoenzyme α subunit. A DnaE single-enzyme assay was exquisitely sensitive to inhibition by nargenicin, and other in vitro characterization studies corroborated DnaE as the target. Medicinal chemistry efforts may expand the spectrum of this novel mechanism antibiotic.
authors.affiliation: In vitro Pharmacology, Merck Research Laboratories, Kenilworth, NJ 07033, USA.
authors.forename: Ronald E
authors.initials: RE
authors.lastname: Painter
authors.affiliation: Screening and Protein Sciences, Merck Research Laboratories, North Wales, PA 19454, USA.
authors.forename: Gregory C
authors.initials: GC
authors.lastname: Adam
authors.affiliation: Medicinal Chemistry, Merck Research Laboratories, Kenilworth, NJ 07033, USA.
authors.forename: Marta
authors.initials: M
authors.lastname: Arocho
authors.affiliation: In vitro Pharmacology, Merck Research Laboratories, Kenilworth, NJ 07033, USA.
authors.forename: Edward
authors.initials: E
authors.lastname: DiNunzio
authors.affiliation: Infectious Disease Biology, Merck Research Laboratories, Kenilworth, NJ 07033, USA.
authors.forename: Robert G K
authors.initials: RG
authors.lastname: Donald
authors.affiliation: Infectious Disease Biology, Merck Research Laboratories, Kenilworth, NJ 07033, USA.
authors.forename: Karen
authors.initials: K
authors.lastname: Dorso
authors.affiliation: Centro de Investigación Básica (CIBE), Merck Sharp & Dhome de España, S.A., 28027 Madrid, Spain.
authors.forename: Olga
authors.initials: O
authors.lastname: Genilloud
authors.affiliation: Infectious Disease Biology, Merck Research Laboratories, Kenilworth, NJ 07033, USA.
authors.forename: Charles
authors.initials: C
authors.lastname: Gill
authors.affiliation: Medicinal Chemistry, Merck Research Laboratories, Kenilworth, NJ 07033, USA.
authors.forename: Michael
authors.initials: M
authors.lastname: Goetz
authors.affiliation: Infectious Disease Biology, Merck Research Laboratories, Kenilworth, NJ 07033, USA.
authors.forename: Nichelle N
authors.initials: NN
authors.lastname: Hairston
authors.affiliation: Discovery Pharmacogenomics, Merck Research Laboratories, Kenilworth, NJ 07033, USA.
authors.forename: Nicholas
authors.initials: N
authors.lastname: Murgolo
authors.affiliation: Infectious Disease Biology, Merck Research Laboratories, Kenilworth, NJ 07033, USA.
authors.forename: Bakela
authors.initials: B
authors.lastname: Nare
authors.affiliation: Infectious Disease Biology, Merck Research Laboratories, Kenilworth, NJ 07033, USA.
authors.forename: David B
authors.initials: DB
authors.lastname: Olsen
authors.affiliation: Infectious Disease Biology, Merck Research Laboratories, Kenilworth, NJ 07033, USA.
authors.forename: Maryann
authors.initials: M
authors.lastname: Powles
authors.affiliation: Infectious Disease Biology, Merck Research Laboratories, Kenilworth, NJ 07033, USA.
authors.forename: Fred
authors.initials: F
authors.lastname: Racine
authors.affiliation: Medicinal Chemistry, Merck Research Laboratories, Kenilworth, NJ 07033, USA.
authors.forename: Jing
authors.initials: J
authors.lastname: Su
authors.affiliation: Centro de Investigación Básica (CIBE), Merck Sharp & Dhome de España, S.A., 28027 Madrid, Spain.
authors.forename: Francisca
authors.initials: F
authors.lastname: Vicente
authors.affiliation: Infectious Disease Biology, Merck Research Laboratories, Kenilworth, NJ 07033, USA.
authors.forename: Douglas
authors.initials: D
authors.lastname: Wisniewski
authors.affiliation: Medicinal Chemistry, Merck Research Laboratories, Kenilworth, NJ 07033, USA.
authors.forename: Li
authors.initials: L
authors.lastname: Xiao
authors.affiliation: Infectious Disease Biology, Merck Research Laboratories, Kenilworth, NJ 07033, USA.
authors.forename: Milton
authors.initials: M
authors.lastname: Hammond
authors.affiliation: Infectious Disease Biology, Merck Research Laboratories, Kenilworth, NJ 07033, USA. Electronic address: katherine_young@merck.com.
authors.forename: Katherine
authors.initials: K
authors.lastname: Young
mesh_terms: D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D004258:DNA Polymerase III / Q000235:genetics*; D004261:DNA Replication / Q000187:drug effects; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D055808:Drug Discovery; D024881:Drug Resistance, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000187:drug effects; D020128:Inhibitory Concentration 50; D007783:Lactones / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D009154:Mutation; D019384:Nucleic Acid Synthesis Inhibitors / Q000737:chemistry / Q000494:pharmacology; D013211:Staphylococcus aureus / Q000187:drug effects
chemical_list: D000900:Anti-Bacterial Agents; D007783:Lactones; D019384:Nucleic Acid Synthesis Inhibitors; C034624:nargenicin A1; C074904:DNA polymerase III, alpha subunit; D004258:DNA Polymerase III; D004259:DNA-Directed DNA Polymerase