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Biophys J. 2014 Jul 1;107(1):185-96. doi: 10.1016/j.bpj.2014.05.025.

Probing the conformation of FhaC with small-angle neutron scattering and molecular modeling.

Author information

1
Université Grenoble Alpes, IBS, F-38044 Grenoble, France; CNRS, IBS, F-38044 Grenoble, France; CEA, IBS, F-38044 Grenoble, France; Institut Laue-Langevin, Grenoble, France. Electronic address: frank.gabel@ibs.fr.
2
CNRS USR 3078, Institut de Recherche Interdisciplinaire, Campus CNRS de la Haute Borne, Université Lille Nord de France, IFR 147, BP 70478, Villeneuve d'Ascq, France. Electronic address: marc.lensink@iri.univ-lille1.fr.
3
CNRS USR 3078, Institut de Recherche Interdisciplinaire, Campus CNRS de la Haute Borne, Université Lille Nord de France, IFR 147, BP 70478, Villeneuve d'Ascq, France.
4
Institut Pasteur de Lille, Centre d'Infection et d'Immunité de Lille, Lille, France; CNRS UMR8204, Lille, France; INSERM U1019, Lille, France; Université Lille Nord de France, Lille, France.
5
Université Grenoble Alpes, IBS, F-38044 Grenoble, France; CNRS, IBS, F-38044 Grenoble, France; CEA, IBS, F-38044 Grenoble, France. Electronic address: christine.ebel@ibs.fr.

Abstract

Probing the solution structure of membrane proteins represents a formidable challenge, particularly when using small-angle scattering. Detergent molecules often present residual scattering contributions even at their match point in small-angle neutron scattering (SANS) measurements. Here, we studied the conformation of FhaC, the outer-membrane, β-barrel transporter of the Bordetella pertussis filamentous hemagglutinin adhesin. SANS measurements were performed on homogeneous solutions of FhaC solubilized in n-octyl-d17-βD-glucoside and on a variant devoid of the α helix H1, which critically obstructs the FhaC pore, in two solvent conditions corresponding to the match points of the protein and the detergent, respectively. Protein-bound detergent amounted to 142 ± 10 mol/mol as determined by analytical ultracentrifugation. By using molecular modeling and starting from three distinct conformations of FhaC and its variant embedded in lipid bilayers, we generated ensembles of protein-detergent arrangement models with 120-160 detergent molecules. The scattered curves were back-calculated for each model and compared with experimental data. Good fits were obtained for relatively compact, connected detergent belts, which occasionally displayed small detergent-free patches on the outer surface of the β barrel. The combination of SANS and modeling clearly enabled us to infer the solution structure of FhaC, with H1 inside the pore as in the crystal structure. We believe that our strategy of combining explicit atomic detergent modeling with SANS measurements has significant potential for structural studies of other detergent-solubilized membrane proteins.

PMID:
24988353
PMCID:
PMC4119272
DOI:
10.1016/j.bpj.2014.05.025
[Indexed for MEDLINE]
Free PMC Article

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