title: Solution structure of 4'-phosphopantetheine - GmACP3 from Geobacter metallireducens: a specialized acyl carrier protein with atypical structural features and a putative role in lipopolysaccharide biosynthesis.
abstract: GmACP3 from Geobacter metallireducens is a specialized acyl carrier protein (ACP) whose gene, gmet_2339, is located near genes encoding many proteins involved in lipopolysaccharide (LPS) biosynthesis, indicating a likely function for GmACP3 in LPS production. By overexpression in Escherichia coli, about 50% holo-GmACP3 and 50% apo-GmACP3 were obtained. Apo-GmACP3 exhibited slow precipitation and non-monomeric behavior by (15)N NMR relaxation measurements. Addition of 4'-phosphopantetheine (4'-PP) via enzymatic conversion by E. coli holo-ACP synthase resulted in stable >95% holo-GmACP3 that was characterized as monomeric by (15)N relaxation measurements and had no indication of conformational exchange. We have determined a high-resolution solution structure of holo-GmACP3 by standard NMR methods, including refinement with two sets of NH residual dipolar couplings, allowing for a detailed structural analysis of the interactions between 4'-PP and GmACP3. Whereas the overall four helix bundle topology is similar to previously solved ACP structures, this structure has unique characteristics, including an ordered 4'-PP conformation that places the thiol at the entrance to a central hydrophobic cavity near a conserved hydrogen-bonded Trp-His pair. These residues are part of a conserved WDSLxH/N motif found in GmACP3 and its orthologs. The helix locations and the large hydrophobic cavity are more similar to medium- and long-chain acyl-ACPs than to other apo- and holo-ACP structures. Taken together, structural characterization along with bioinformatic analysis of nearby genes suggests that GmACP3 is involved in lipid A acylation, possibly by atypical long-chain hydroxy fatty acids, and potentially is involved in synthesis of secondary metabolites.
authors.affiliation: Department of Chemistry and Biochemistry, Miami University, Oxford, Ohio 45056, United States.
authors.forename: Theresa A
authors.initials: TA
authors.lastname: Ramelot
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Smola
authors.forename: Hsiau-Wei
authors.initials: HW
authors.lastname: Lee
authors.forename: Colleen
authors.initials: C
authors.lastname: Ciccosanti
authors.forename: Keith
authors.initials: K
authors.lastname: Hamilton
authors.forename: Thomas B
authors.initials: TB
authors.lastname: Acton
authors.forename: Rong
authors.initials: R
authors.lastname: Xiao
authors.forename: John K
authors.initials: JK
authors.lastname: Everett
authors.forename: James H
authors.initials: JH
authors.lastname: Prestegard
authors.forename: Gaetano T
authors.initials: GT
authors.lastname: Montelione
authors.forename: Michael A
authors.initials: MA
authors.lastname: Kennedy
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli; D015964:Gene Expression Regulation, Bacterial; D045302:Geobacter / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000096:biosynthesis*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010204:Pantetheine / Q000031:analogs & derivatives* / Q000737:chemistry; D011487:Protein Conformation
chemical_list: D001426:Bacterial Proteins; D002352:Carrier Proteins; D008070:Lipopolysaccharides; D010204:Pantetheine; C003129:4'-phosphopantetheine