Conformational transitions in the glycine-bound GluN1 NMDA receptor LBD via single-molecule FRET.

Autor:	Cooper DR; Department of Chemistry, Rice University, Houston, Texas., Dolino DM; Center for Membrane Biology, Department of Biochemistry and Molecular Biology, Graduate School of Biomedical Sciences, University of Texas Health Science Center, Houston, Texas., Jaurich H; Department of Chemistry, Rice University, Houston, Texas., Shuang B; Department of Chemistry, Rice University, Houston, Texas., Ramaswamy S; Center for Membrane Biology, Department of Biochemistry and Molecular Biology, Graduate School of Biomedical Sciences, University of Texas Health Science Center, Houston, Texas., Nurik CE; Center for Membrane Biology, Department of Biochemistry and Molecular Biology, Graduate School of Biomedical Sciences, University of Texas Health Science Center, Houston, Texas., Chen J; Department of Chemistry, Rice University, Houston, Texas., Jayaraman V; Center for Membrane Biology, Department of Biochemistry and Molecular Biology, Graduate School of Biomedical Sciences, University of Texas Health Science Center, Houston, Texas. Electronic address: vasanthi.jayaraman@uth.tmc.edu., Landes CF; Department of Chemistry, Rice University, Houston, Texas; Department of Electrical and Computer Engineering, Rice University, Houston, Texas. Electronic address: cflandes@rice.edu.
Jazyk:	angličtina
Zdroj:	Biophysical journal [Biophys J] 2015 Jul 07; Vol. 109 (1), pp. 66-75.
DOI:	10.1016/j.bpj.2015.05.025
Abstrakt:	The N-methyl-D-aspartate receptor (NMDAR) is a member of the glutamate receptor family of proteins and is responsible for excitatory transmission. Activation of the receptor is thought to be controlled by conformational changes in the ligand binding domain (LBD); however, glutamate receptor LBDs can occupy multiple conformations even in the activated form. This work probes equilibrium transitions among NMDAR LBD conformations by monitoring the distance across the glycine-bound LBD cleft using single-molecule Förster resonance energy transfer (smFRET). Recent improvements in photoprotection solutions allowed us to monitor transitions among the multiple conformations. Also, we applied a recently developed model-free algorithm called "step transition and state identification" to identify the number of states, their smFRET efficiencies, and their interstate kinetics. Reversible interstate conversions, corresponding to transitions among a wide range of cleft widths, were identified in the glycine-bound LBD, on much longer timescales compared to channel opening. These transitions were confirmed to be equilibrium in nature by shifting the distribution reversibly via denaturant. We found that the NMDAR LBD proceeds primarily from one adjacent smFRET state to the next under equilibrium conditions, consistent with a cleft-opening/closing mechanism. Overall, by analyzing the state-to-state transition dynamics and distributions, we achieve insight into specifics of long-lived LBD equilibrium structural dynamics, as well as obtain a more general description of equilibrium folding/unfolding in a conformationally dynamic protein. The relationship between such long-lived LBD dynamics and channel function in the full receptor remains an open and interesting question. (Copyright © 2015 Biophysical Society. Published by Elsevier Inc. All rights reserved.)
Databáze:	MEDLINE
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