AUC and Small-Angle Scattering for Membrane Proteins.

Abstract : Analytical ultracentrifugation is a key tool to assess homogeneity of membrane protein samples, to determine protein association state and detergent concentration, and to characterize protein-protein equilibrium. Combining absorbance and interference detections gives information on the amount of the detergent and lipid bound to proteins. Changing the solvent density affects specifically the buoyancy of each of the different components, and can also be used to gain information on particle composition and interaction. We will present the related tools, recently implemented in the softwares Sedphat (sedfitsedphat.nibib.nih.gov/software) and Gussi (http://biophysics.swmed.edu/MBR/software.html), which help to measure the amount of detergent bound to the protein, and ascertain the protein association state within the protein-detergent complex. In addition, fluorescence detection allows focusing specifically on a labeled component within a complex mixture. We present two examples of sedimentation velocity experiments, allowing on one hand to evidence complex formation between an unpurified GFP-labeled protein and a membrane protein, and on the other hand to characterize fluorescent lipid vesicles. Small-angle X-ray and neutron scattering are techniques that give insights into the structure and conformation of macromolecules in solution. However, the detergents used to purify membrane protein are often imperfectly masked due to their amphipathic character. Particular strategies addressing membrane proteins were recently proposed, which are shortly presented.
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Article dans une revue
Methods in Enzymology, Elsevier, 2015, 562, pp.257-86
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http://hal.univ-grenoble-alpes.fr/hal-01234685
Contributeur : Frank Thomas <>
Soumis le : vendredi 27 novembre 2015 - 11:45:07
Dernière modification le : jeudi 11 janvier 2018 - 06:15:24

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Aline Le Roy, Kai Wang, Béatrice Schaack, Peter Schuck, Cécile Breyton, et al.. AUC and Small-Angle Scattering for Membrane Proteins.. Methods in Enzymology, Elsevier, 2015, 562, pp.257-86. 〈hal-01234685〉

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