The Journal of Membrane Biology

, Volume 247, Issue 9–10, pp 981–996 | Cite as

Solution Behavior and Crystallization of Cytochrome bc1 in the Presence of Amphipols

  • Delphine Charvolin
  • Martin PicardEmail author
  • Li-Shar Huang
  • Edward A. Berry
  • Jean-Luc Popot


Detergents classically are used to keep membrane proteins soluble in aqueous solutions, but they tend to destabilize them. This problem can be largely alleviated thanks to the use of amphipols (APols), small amphipathic polymers designed to substitute for detergents. APols adsorb at the surface of the transmembrane region of membrane proteins, keeping them water-soluble while stabilizing them bio-chemically. Membrane protein/APol complexes have proven, however, difficult to crystallize. In this study, the composition and solution properties of complexes formed between mitochondrial cytochrome bc1 and A8-35, the most extensively used APol to date, have been studied by means of size exclusion chromatography, sucrose gradient sedimentation, and small-angle neutron scattering. Stable, monodisperse preparations of bc1/A8-35 complexes can be obtained, which, depending on the medium, undergo either repulsive or attractive interactions. Under crystallization conditions, diffracting three-dimensional crystals of A8-35-stabilized cytochrome bc1 formed, but only in the concomitant presence of APol and detergent.


Membrane proteins Surfactants Amphipathic polymers Stability Small-angle neutron scattering Size exclusion chromatography 



A specific type of amphipol (Tribet et al. 1996)




Analytical ultracentrifugation




Tetraethylene glycol monooctyl ether


Critical micellar concentration


Contrast match point




Ethylene-diamine-tetra-acetic acid


Electron microscopy


Fluorescent (NBD-labeled) A8-35


2-Morpholino-ethane sulfonic acid potassium salt


Molecular dynamics


Molecular weight


Molecular weight cutoff




Nuclear magnetic resonance




Outer membrane protein X from Escherichia coli


4 kDa polyethylene glycol


Small-angle neutron and X-ray scattering, respectively


Size exclusion chromatography


Sarcoplasmic reticulum calcium pump from fast twitch muscle


Transmembrane domain of outer membrane protein A from E. coli





Particular thanks are due to A.-N. Galatanu and J.-C. Courant for participating in some of these experiments, to P. Hervé for the synthesis of [3H]A8-35, to P. Timmins for his participation in the SANS measurements, to D. Picot and L. Barucq for testing crystals at the ESRF, to I. Gallay for help with the X-ray equipment at the IBPC, to J. Barra and L.J. Catoire for their precious help with the figures, and to Y. Gohon and M. Zoonens for useful comments on the manuscript. This study was supported by the Centre National de la Recherche Scientifique, Université Paris-7, the Human Frontier Science Program Organization (Grant RG00223-2000-M), E.U. Specific Targeted Research Project LSHG-CT-2005-513770 IMPS (Innovative tools for membrane protein structural proteomics), and NIH grant R01DK44842. The stays of EAB in France were subsidized by awards from the France-Berkeley Fund and the French Ministère de l’Education Nationale et de la Recherche.


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Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  • Delphine Charvolin
    • 1
  • Martin Picard
    • 1
    • 2
    Email author
  • Li-Shar Huang
    • 3
  • Edward A. Berry
    • 3
  • Jean-Luc Popot
    • 1
  1. 1.UMR 7099, Centre National de la Recherche Scientifique/Université Paris-7, Institut de Biologie Physico-Chimique, FRC 550ParisFrance
  2. 2.Faculté de Pharmacie, Laboratoire de Cristallographie et RMN BiologiquesCentre National de la Recherche Scientifique/Université Paris Descartes UMR 8015ParisFrance
  3. 3.Department of Biochemistry and Molecular BiologySUNY Upstate Medical UniversitySyracuseUSA

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