Strategy of macromolecular grafting onto a gold substrate dedicated to protein-protein interaction measurements.

Virginie Mansuy-Schlick (1) , Régis Delage-Mourroux (1) , Michèle Jouvenot (1) , Wilfrid Boireau (2)

1 E2SNC / ESTROGENES - Estrogènes, Expression génique et pathologies du Système Nerveux Central - UFC
2 FEMTO-ST - Franche-Comté Électronique Mécanique, Thermique et Optique - Sciences et Technologies (UMR 6174)

Many biotechnology applications use proteins immobilized on surface. For biosensor, the sensing layer is a key component interfacing the transducer and the sample. Strategies employed to activate the bidimensional surface act directly on the performance of the biosensor. In this paper we propose a novel strategy for engineered proteins self-assembly. Our original supramolecular structure allows a direct and fast covalent attachment of proteins onto bare gold substrate through a homobifunctional cross-linker, 1,4-di-([2'-pyridyldithio]propionamido)butane (DPDPB). In this work, engineered proteins and linker-protein complexes were synthesized and characterized by gel electrophoresis, chromatography and spectroscopy experiments. Macromolecular construction "DPDPB-GST tag-GEC1 protein" was conceived in order to guarantee a 2D architecture enhancing the capabilities of the target (tubulin) to recognize its partner (GEC1). Surface plasmon resonance measurements clearly showed potential of this particular self-assembled protein layer compared to a commercial immunosensor interface. At the concentrations tested, the recognition process occurs between tubulin and the immobilized GEC1; moreover enhanced binding was obtained with the home-made 2D sensing layer more than with 3D carboxymethyl dextran matrix.

Mots clés: Self-assembled monolayer (SAM); DPDPB; Protein immobilization; Surface plasmon resonance (SPR); GEC1; Tubulin

Domaines: Biophysique

Virginie Mansuy-Schlick, Régis Delage-Mourroux, Michèle Jouvenot, Wilfrid Boireau. Strategy of macromolecular grafting onto a gold substrate dedicated to protein-protein interaction measurements.. Biosensors and Bioelectronics, 2006, 21 (9), pp.1830-7. ⟨10.1016/j.bios.2005.11.021⟩. ⟨hal-00446991⟩