title: Crystallization and X-ray diffraction analysis of the complement component-3 (C3) inhibitory domain of Efb from Staphylococcus aureus.
abstract: The extracellular fibrinogen-binding protein (Efb) of Staphylococcus aureus is a multifunctional virulence factor capable of potent inhibition of complement component-3 (C3) activity in addition to its previously described fibrinogen-binding properties. A truncated recombinant form of Efb (Efb-C) that binds C3 has been overexpressed and purified and has been crystallized using the hanging-drop vapor-diffusion technique. Crystals of native Efb-C grew in the tetragonal space group P4(3) (unit-cell parameters a = b = 59.53, c = 46.63 A) with two molecules in the asymmetric unit and diffracted well beyond 1.25 A limiting Bragg spacing. To facilitate de novo phasing of the Efb-C crystals, two independent site-directed mutants were engineered in which either residue Ile112 or Val140 was replaced with methionine and crystals isomorphous to those of native Efb-C were reproduced using a seleno-L-methionine-labeled form of each mutant protein. Multiwavelength anomalous diffraction (MAD) data were collected on both mutants and analyzed for their phasing power toward solution and refinement of a high-resolution Efb-C crystal structure.
authors.affiliation: Division of Cell Biology and Biophysics, School of Biological Sciences, University of Missouri-Kansas City, USA.
authors.forename: Michal
authors.initials: M
authors.lastname: Hammel
authors.forename: Kasra X
authors.initials: KX
authors.lastname: Ramyar
authors.forename: Charles T
authors.initials: CT
authors.lastname: Spencer
authors.forename: Brian V
authors.initials: BV
authors.lastname: Geisbrecht
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D003176:Complement C3 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D003460:Crystallization / Q000379:methods; D018360:Crystallography, X-Ray; D016297:Mutagenesis, Site-Directed; D017434:Protein Structure, Tertiary / Q000235:genetics; D013211:Staphylococcus aureus / Q000737:chemistry*
chemical_list: D001426:Bacterial Proteins; D003176:Complement C3; C114014:Efb protein, Staphylococcus aureus