Single-Molecule Patch-Clamp FRET Anisotropy Microscopy Studies of NMDA Receptor Ion Channel Activation and Deactivation under Agonist Ligand Binding in Living Cells

• DOI: 10.1021/jacs.6b03496
• Publication Date: 2016-06-07T18:23:48Z
• Authors: Dibyendu Kumar Sa...; Rajeev Yadav; H. Peter Lu
• Keywords: Single-molecule FRET

N-methyl-d-aspartate (NMDA) receptor ion channel is activated by the binding of two pairs glycine and glutamate along with application action potential. Binding unbinding ligands changes its conformation that plays a critical role in open–close activities NMDA receptor. Conformation states their dynamics due to ligand are extremely difficult characterize either conventional ensemble experiments or single-channel electrophysiology method. Here we report development new correlated technical approach, single-molecule patch-clamp FRET anisotropy imaging demonstrate probing kinetics domain. Experimentally determined further verified computational modeling. Single-channel four-channel fluorescence measurement recorded simultaneously get correlation among electrical on off states, optically conformational open closed FRET, binding-unbinding at domain GluN1 subunit. This method has ability detect intermediate addition states. Based our experimental results, have proposed gating goes through least one electrically state, desensitized when remain bound similar fully state.