| Autor: |
Giusti F; Laboratoire de Biologie Physico-Chimique des Protéines Membranaires, UMR 7099, CNRS/Université Paris 7, Institut de Biologie Physico-Chimique (FRC 550), 13 rue Pierre et Marie Curie, F-75005 Paris, France., Kessler P; CEA, Institut de Biologie et de Technologies de Saclay, Service d'Ingénierie Moléculaire des Protéines, 91191 Gif-sur-Yvette, France., Hansen RW; Department of Chemistry & Nano-Science Center, University of Copenhagen , Universitetsparken 5, DK-2100 Copenhagen Ø, Denmark., Della Pia EA; Department of Chemistry & Nano-Science Center, University of Copenhagen , Universitetsparken 5, DK-2100 Copenhagen Ø, Denmark., Le Bon C; Laboratoire de Biologie Physico-Chimique des Protéines Membranaires, UMR 7099, CNRS/Université Paris 7, Institut de Biologie Physico-Chimique (FRC 550), 13 rue Pierre et Marie Curie, F-75005 Paris, France., Mourier G; CEA, Institut de Biologie et de Technologies de Saclay, Service d'Ingénierie Moléculaire des Protéines, 91191 Gif-sur-Yvette, France., Popot JL; Laboratoire de Biologie Physico-Chimique des Protéines Membranaires, UMR 7099, CNRS/Université Paris 7, Institut de Biologie Physico-Chimique (FRC 550), 13 rue Pierre et Marie Curie, F-75005 Paris, France., Martinez KL; Department of Chemistry & Nano-Science Center, University of Copenhagen , Universitetsparken 5, DK-2100 Copenhagen Ø, Denmark., Zoonens M; Laboratoire de Biologie Physico-Chimique des Protéines Membranaires, UMR 7099, CNRS/Université Paris 7, Institut de Biologie Physico-Chimique (FRC 550), 13 rue Pierre et Marie Curie, F-75005 Paris, France. |
| Abstrakt: |
Amphipols (APols) are short amphipathic polymers that stabilize membrane proteins (MPs) in aqueous solutions. In the present study, A8-35, a polyacrylate-based APol, was grafted with hexahistidine tags (His6-tags). The synthesis and characterization of this novel functionalized APol, named HistAPol, are described. Its ability to immobilize MPs on nickel ion-bearing surfaces was tested using two complementary methods, immobilized metal affinity chromatography (IMAC) and surface plasmon resonance (SPR). Compared to a single His6-tag fused at one extremity of a MP, the presence of several His6-tags carried by the APol belt surrounding the transmembrane domain of a MP increases remarkably the affinity of the protein/APol complex for nickel ion-bearing SPR chips, whereas it does not show such a strong effect on an IMAC resin. HistAPol-mediated immobilization, which allows reversibility of the interaction and easy regeneration of the supports and dispenses with any genetic modification of the target protein, provides a novel, promising tool for attaching MPs onto solid supports while stabilizing them. |
