title: Discovery of FabH/FabF inhibitors from natural products.
abstract: Condensing enzymes are essential in type II fatty acid synthesis and are promising targets for antibacterial drug discovery. Recently, a new approach using a xylose-inducible plasmid to express antisense RNA in Staphylococcus aureus has been described; however, the actual mechanism was not delineated. In this paper, the mechanism of decreased target protein production by expression of antisense RNA was investigated using Northern blotting. This revealed that the antisense RNA acts posttranscriptionally by targeting mRNA, leading to 5' mRNA degradation. Using this technology, a two-plate assay was developed in order to identify FabF/FabH target-specific cell-permeable inhibitors by screening of natural product extracts. Over 250,000 natural product fermentation broths were screened and then confirmed in biochemical assays, yielding a hit rate of 0.1%. All known natural product FabH and FabF inhibitors, including cerulenin, thiolactomycin, thiotetromycin, and Tü3010, were discovered using this whole-cell mechanism-based screening approach. Phomallenic acids, which are new inhibitors of FabF, were also discovered. These new inhibitors exhibited target selectivity in the gel elongation assay and in the whole-cell-based two-plate assay. Phomallenic acid C showed good antibacterial activity, about 20-fold better than that of thiolactomycin and cerulenin, against S. aureus. It exhibited a spectrum of antibacterial activity against clinically important pathogens including methicillin-resistant Staphylococcus aureus, Bacillus subtilis, and Haemophilus influenzae.
authors.affiliation: Merck Research Laboratories, Rahway, NJ 07065, USA.
authors.forename: Katherine
authors.initials: K
authors.lastname: Young
authors.forename: Hiranthi
authors.initials: H
authors.lastname: Jayasuriya
authors.forename: John G
authors.initials: JG
authors.lastname: Ondeyka
authors.forename: Kithsiri
authors.initials: K
authors.lastname: Herath
authors.forename: Chaowei
authors.initials: C
authors.lastname: Zhang
authors.forename: Srinivas
authors.initials: S
authors.lastname: Kodali
authors.forename: Andrew
authors.initials: A
authors.lastname: Galgoci
authors.forename: Ronald
authors.initials: R
authors.lastname: Painter
authors.forename: Vickie
authors.initials: V
authors.lastname: Brown-Driver
authors.forename: Robert
authors.initials: R
authors.lastname: Yamamoto
authors.forename: Lynn L
authors.initials: LL
authors.lastname: Silver
authors.forename: Yingcong
authors.initials: Y
authors.lastname: Zheng
authors.forename: Judith I
authors.initials: JI
authors.lastname: Ventura
authors.forename: Janet
authors.initials: J
authors.lastname: Sigmund
authors.forename: Sookhee
authors.initials: S
authors.lastname: Ha
authors.forename: Angela
authors.initials: A
authors.lastname: Basilio
authors.forename: Francisca
authors.initials: F
authors.lastname: Vicente
authors.forename: José Rubén
authors.initials: JR
authors.lastname: Tormo
authors.forename: Fernando
authors.initials: F
authors.lastname: Pelaez
authors.forename: Phil
authors.initials: P
authors.lastname: Youngman
authors.forename: Doris
authors.initials: D
authors.lastname: Cully
authors.forename: John F
authors.initials: JF
authors.lastname: Barrett
authors.forename: Dennis
authors.initials: D
authors.lastname: Schmatz
authors.forename: Sheo B
authors.initials: SB
authors.lastname: Singh
authors.forename: Jun
authors.initials: J
authors.lastname: Wang
mesh_terms: D015099:3-Oxoacyl-(Acyl-Carrier-Protein) Synthase / Q000037:antagonists & inhibitors*; D000900:Anti-Bacterial Agents / Q000302:isolation & purification / Q000494:pharmacology*; D001419:Bacteria / Q000201:enzymology*; D001688:Biological Products / Q000737:chemistry*; D015195:Drug Design; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005227:Fatty Acids / Q000096:biosynthesis; D008826:Microbial Sensitivity Tests; D016372:RNA, Antisense / Q000494:pharmacology; D012333:RNA, Messenger / Q000737:chemistry; D013329:Structure-Activity Relationship
chemical_list: D000900:Anti-Bacterial Agents; D001688:Biological Products; D004791:Enzyme Inhibitors; D005227:Fatty Acids; D016372:RNA, Antisense; D012333:RNA, Messenger; C077402:3-ketoacyl-acyl carrier protein synthase III; D015099:3-Oxoacyl-(Acyl-Carrier-Protein) Synthase