The Journal of Membrane Biology

, Volume 247, Issue 9–10, pp 925–940 | Cite as

Amphipol-Mediated Screening of Molecular Orthoses Specific for Membrane Protein Targets

  • Yann Ferrandez
  • Manuela Dezi
  • Mickael Bosco
  • Agathe Urvoas
  • Marie Valerio-Lepiniec
  • Christel Le Bon
  • Fabrice Giusti
  • Isabelle Broutin
  • Grégory Durand
  • Ange Polidori
  • Jean-Luc Popot
  • Martin PicardEmail author
  • Philippe Minard


Specific, tight-binding protein partners are valuable helpers to facilitate membrane protein (MP) crystallization, because they can i) stabilize the protein, ii) reduce its conformational heterogeneity, and iii) increase the polar surface from which well-ordered crystals can grow. The design and production of a new family of synthetic scaffolds (dubbed αReps, for “artificial alpha repeat protein”) have been recently described. The stabilization and immobilization of MPs in a functional state are an absolute prerequisite for the screening of binders that recognize specifically their native conformation. We present here a general procedure for the selection of αReps specific of any MP. It relies on the use of biotinylated amphipols, which act as a universal “Velcro” to stabilize, and immobilize MP targets onto streptavidin-coated solid supports, thus doing away with the need to tag the protein itself.


HEAT repeat protein Protein design Phage display Membrane protein Amphipols Immobilization 



E. coli rich media


A specific type of poly(acrylic acid)-based amphipol




Biotinylated A8-35


Biotinylated non-ionic amphipol




Critical micellar concentration




Number-average degree of polymerization


Ethylene diamine tetraacetic acid


Electron microscopy


Hexahistidine tag




Molecular dynamics


Number-average molar mass


Membrane protein


Molecular weight


MW cut-off


Non-ionic amphipol


Nuclear magnetic resonance


Nitrilotriacetic acid


Optical density measured at 600 nm




Phosphate buffer saline


Polyethylene glycol


Sodium dodecyl sulfate polyacrylamide gel electrophoresis


Size-exclusion chromatography


Tris-buffered saline


Tris-buffered saline supplemented with Tween 20 (w/v)





Particular thanks are due to J.D. Perlmutter and J.N. Sachs for communication of a molecular dynamics model of an A8-35 particle. This research was supported by ANR-2010-BLAN-1535, by the French Centre National de la Recherche Scientifique (CNRS), by Université Paris-5 Paris Descartes, by Université Paris-7 Denis Diderot, and by grant “DYNAMO”, ANR-11-LABX-0011-01 from the French “Initiative d’Excellence” program.


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Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  • Yann Ferrandez
    • 1
  • Manuela Dezi
    • 2
  • Mickael Bosco
    • 3
    • 4
  • Agathe Urvoas
    • 1
  • Marie Valerio-Lepiniec
    • 1
  • Christel Le Bon
    • 5
  • Fabrice Giusti
    • 5
  • Isabelle Broutin
    • 2
  • Grégory Durand
    • 3
    • 4
  • Ange Polidori
    • 3
    • 4
  • Jean-Luc Popot
    • 5
  • Martin Picard
    • 2
    Email author
  • Philippe Minard
    • 1
  1. 1.Laboratoire de Modélisation et Ingénierie des ProtéinesIBBMC UMR 8619, CNRS/Université Paris SudOrsayFrance
  2. 2.Laboratoire de Cristallographie et RMN Biologiques, Faculté de Pharmacie, UMR 8015CNRS/Université Paris DescartesParis Cedex 06France
  3. 3.Equipe Chimie Bioorganique et Systèmes AmphiphilesUniversité d’AvignonAvignonFrance
  4. 4.Institut des Biomolécules Max Mousseron (UMR 5247)Montpellier Cedex 05France
  5. 5.Laboratoire de Biologie Physico-Chimique des Protéines Membranaires, UMR 7099Institut de Biologie Physico-Chimique (FRC 550), Centre National de la Recherche Scientifique/Université Paris-7ParisFrance

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