title: Apo, Zn2+-bound and Mn2+-bound structures reveal ligand-binding properties of SitA from the pathogen Staphylococcus pseudintermedius.
abstract: The Gram-positive bacterium Staphylococcus pseudintermedius is a leading cause of canine bacterial pyoderma, resulting in worldwide morbidity in dogs. S. pseudintermedius also causes life-threatening human infections. Furthermore, methicillin-resistant S. pseudintermedius is emerging, resembling the human health threat of methicillin-resistant Staphylococcus aureus. Therefore it is increasingly important to characterize targets for intervention strategies to counteract S. pseudintermedius infections. Here we used biophysical methods, mutagenesis, and X-ray crystallography, to define the ligand-binding properties and structure of SitA, an S. pseudintermedius surface lipoprotein. SitA was strongly and specifically stabilized by Mn2+ and Zn2+ ions. Crystal structures of SitA complexed with Mn2+ and Zn2+ revealed a canonical class III solute-binding protein with the metal cation bound in a cavity between N- and C-terminal lobes. Unexpectedly, one crystal contained both apo- and holo-forms of SitA, revealing a large side-chain reorientation of His64, and associated structural differences accompanying ligand binding. Such conformational changes may regulate fruitful engagement of the cognate ABC (ATP-binding cassette) transporter system (SitBC) required for metal uptake. These results provide the first detailed characterization and mechanistic insights for a potential therapeutic target of the major canine pathogen S. pseudintermedius, and also shed light on homologous structures in related staphylococcal pathogens afflicting humans.
authors.affiliation: *Novartis Vaccines and Diagnostics srl, Via Fiorentina 1, 53100 Siena, Italy.
authors.forename: Francesca
authors.initials: F
authors.lastname: Abate
authors.affiliation: *Novartis Vaccines and Diagnostics srl, Via Fiorentina 1, 53100 Siena, Italy.
authors.forename: Enrico
authors.initials: E
authors.lastname: Malito
authors.affiliation: *Novartis Vaccines and Diagnostics srl, Via Fiorentina 1, 53100 Siena, Italy.
authors.forename: Roberta
authors.initials: R
authors.lastname: Cozzi
authors.affiliation: *Novartis Vaccines and Diagnostics srl, Via Fiorentina 1, 53100 Siena, Italy.
authors.forename: Paola
authors.initials: P
authors.lastname: Lo Surdo
authors.affiliation: *Novartis Vaccines and Diagnostics srl, Via Fiorentina 1, 53100 Siena, Italy.
authors.forename: Domenico
authors.initials: D
authors.lastname: Maione
authors.affiliation: *Novartis Vaccines and Diagnostics srl, Via Fiorentina 1, 53100 Siena, Italy.
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Bottomley
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001053:Apolipoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002152:Calorimetry, Differential Scanning; D002942:Circular Dichroism; D018360:Crystallography, X-Ray; D004283:Dog Diseases / Q000382:microbiology; D004285:Dogs; D006801:Humans; D008345:Manganese / Q000737:chemistry / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D020125:Mutation, Missense; D011485:Protein Binding; D055503:Protein Multimerization; D055550:Protein Stability; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D013203:Staphylococcal Infections / Q000382:microbiology; D013210:Staphylococcus / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D015032:Zinc / Q000737:chemistry / Q000378:metabolism*
chemical_list: D001053:Apolipoproteins; D001426:Bacterial Proteins; D008345:Manganese; D015032:Zinc