Open AccessCrystallization and X‐ray diffraction analysis of the complement component‐3 (C3) inhibitory domain of Efb from Staphylococcus aureus
Author(s) -
Hammel Michal,
Ramyar Kasra X.,
Spencer Charles T.,
Geisbrecht Brian V.
Publication year - 2006
Publication title -
acta crystallographica section f
Language(s) - English
Resource type - Journals
ISSN - 1744-3091
DOI - 10.1107/s1744309106005926
Subject(s) - complement (music) , staphylococcus aureus , crystallization , diffraction , x ray crystallography , x ray , inhibitory postsynaptic potential , domain (mathematical analysis) , component (thermodynamics) , materials science , chemistry , crystallography , microbiology and biotechnology , physics , bacteria , biology , optics , mathematics , biochemistry , organic chemistry , gene , mathematical analysis , genetics , neuroscience , complementation , phenotype , thermodynamics
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 P 4 3 (unit‐cell parameters a = b = 59.53, c = 46.63 Å) with two molecules in the asymmetric unit and diffracted well beyond 1.25 Å 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.