Two-point immobilization of a conformation-specific beta2-adrenoceptor for recognizing the receptor agonists or antagonists inspired by binding-induced DNA assembly.

Immobilized protein has advanced many areas like drug discovery. While this field evolved rapidly over the three decades, the immobilization platform for G-protein-coupled receptor (GPCR) remains unpromising due to its instability under the relatively harsh conditions of current methodologies. Taking beta2-adrenoceptor (β2-AR) as an example, we presented here a general strategy for immobilization of GPCRs by combing his-tag trap system, conformation-specific aptamer, and target binding induced DNA hybridization. Morphology characterization by diverse assays confirmed a monolayer of β2-AR on the microsphere surface. Radio-ligand binding assay and immuno-transmission electron microscopy displayed desirable ligand- and antibody-binding activities. Owing to the competitive strand displacement during the immobilization, the method proved to be capable of sensitively and directly determining the receptor density on the surface which enormously challenges most of the reported assays. A case study of chromatography using the immobilized receptor as stationary phase exhibited a demonstrable conformation specificity that enables the selective recognition of the receptor agonists or antagonists. This method is possible to turn into a general strategy for immobilization of GPCRs with defined orientation, conformation, function, and density, thus paving a way to precisely realize the receptor-ligand binding interaction and screen the receptor agonist or antagonist with high efficiency.