title: Structure of the CFA/III major pilin subunit CofA from human enterotoxigenic Escherichia coli determined at 0.90 Å resolution by sulfur-SAD phasing.
abstract: CofA, a major pilin subunit of colonization factor antigen III (CFA/III), forms pili that mediate small-intestinal colonization by enterotoxigenic Escherichia coli (ETEC). In this study, the crystal structure of an N-terminally truncated version of CofA was determined by single-wavelength anomalous diffraction (SAD) phasing using five sulfurs in the protein. Given the counterbalance between anomalous signal strength and the undesired X-ray absorption of the solvent, diffraction data were collected at 1.5 Å resolution using synchrotron radiation. These data were sufficient to elucidate the sulfur substructure at 1.38 Å resolution. The low solvent content (29%) of the crystal necessitated that density modification be performed with an additional 0.9 Å resolution data set to reduce the phase error caused by the small sulfur anomalous signal. The CofA structure showed the αβ-fold typical of type IVb pilins and showed high structural homology to that of TcpA for toxin-coregulated pili of Vibrio cholerae, including spatial distribution of key residues critical for pilin self-assembly. A pilus-filament model of CofA was built by computational docking and molecular-dynamics simulation using the previously reported filament model of TcpA as a structural template. This model revealed that the CofA filament surface was highly negatively charged and that a 23-residue-long loop between the α1 and α2 helices filled the gap between the pilin subunits. These characteristics could provide a unique binding epitope for the CFA/III pili of ETEC compared with other type IVb pili.
authors.affiliation: Department of Pharmaceutical Sciences, Osaka University, 1-6 Yamadaoka, Suita, Osaka 565-0871, Japan.
authors.forename: Shunsuke
authors.initials: S
authors.lastname: Fukakusa
authors.forename: Kazuki
authors.initials: K
authors.lastname: Kawahara
authors.forename: Shota
authors.initials: S
authors.lastname: Nakamura
authors.forename: Takaki
authors.initials: T
authors.lastname: Iwashita
authors.forename: Seiki
authors.initials: S
authors.lastname: Baba
authors.forename: Mitsuhiro
authors.initials: M
authors.lastname: Nishimura
authors.forename: Yuji
authors.initials: Y
authors.lastname: Kobayashi
authors.forename: Takeshi
authors.initials: T
authors.lastname: Honda
authors.forename: Tetsuya
authors.initials: T
authors.lastname: Iida
authors.forename: Tooru
authors.initials: T
authors.lastname: Taniguchi
authors.forename: Tadayasu
authors.initials: T
authors.lastname: Ohkubo
mesh_terms: D018360:Crystallography, X-Ray; D054307:Enterotoxigenic Escherichia coli / Q000737:chemistry*; D029968:Escherichia coli Proteins / Q000737:chemistry*; D036781:Fimbriae Proteins / Q000737:chemistry* / Q000145:classification; D010861:Fimbriae, Bacterial / Q000737:chemistry*; D006801:Humans; D021122:Protein Subunits / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D013455:Sulfur / Q000737:chemistry*
chemical_list: C577578:CofA protein, E coli; D029968:Escherichia coli Proteins; D021122:Protein Subunits; C022924:colonization factor antigens; D036781:Fimbriae Proteins; D013455:Sulfur