title: Identification and analysis of the acyl carrier protein (ACP) docking site on beta-ketoacyl-ACP synthase III.
abstract: The molecular details that govern the specific interactions between acyl carrier protein (ACP) and the enzymes of fatty acid biosynthesis are unknown. We investigated the mechanism of ACP-protein interactions using a computational analysis to dock the NMR structure of ACP with the crystal structure of beta-ketoacyl-ACP synthase III (FabH) and experimentally tested the model by the biochemical analysis of FabH mutants. The activities of the mutants were assessed using both an ACP-dependent and an ACP-independent assay. The ACP interaction surface was defined by mutations that compromised FabH activity in the ACP-dependent assay but had no effect in the ACP-independent assay. ACP docked to a positively charged/hydrophobic patch adjacent to the active site tunnel on FabH, which included a conserved arginine (Arg-249) that was required for ACP docking. Kinetic analysis and direct binding studies between FabH and ACP confirmed the identification of Arg-249 as critical for FabH-ACP interaction. Our experiments reveal the significance of the positively charged/hydrophobic patch located adjacent to the active site cavities of the fatty acid biosynthesis enzymes and the high degree of sequence conservation in helix II of ACP across species.
authors.affiliation: Department of Biochemistry, St. Jude Children's Research Hospital, Memphis, Tennessee 38105, USA.
authors.forename: Y M
authors.initials: YM
authors.lastname: Zhang
authors.forename: M S
authors.initials: MS
authors.lastname: Rao
authors.forename: R J
authors.initials: RJ
authors.lastname: Heath
authors.forename: A C
authors.initials: AC
authors.lastname: Price
authors.forename: A J
authors.initials: AJ
authors.lastname: Olson
authors.forename: C O
authors.initials: CO
authors.lastname: Rock
authors.forename: S W
authors.initials: SW
authors.lastname: White
mesh_terms: D015099:3-Oxoacyl-(Acyl-Carrier-Protein) Synthase / Q000737:chemistry* / Q000378:metabolism; D000213:Acyl Carrier Protein / Q000378:metabolism*; D000595:Amino Acid Sequence; D001665:Binding Sites; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D017433:Protein Structure, Secondary
chemical_list: D000213:Acyl Carrier Protein; C077402:3-ketoacyl-acyl carrier protein synthase III; D015099:3-Oxoacyl-(Acyl-Carrier-Protein) Synthase