Structural Mobility of the Extracellular Ligand-Binding Core of an Ionotropic Glutamate Receptor. Analysis of NMR Relaxation Dynamics<sup>†</sup>

2002-07-26T00:00:00Z (GMT) by Robert L. McFeeters Robert E. Oswald
Ionotropic glutamate receptors play important roles in a variety of neuronal processes and have been implicated in multiple neurodegenerative diseases. The extracellular ligand-binding (S1S2) core of the GluR2 subtype can be expressed in bacteria as a soluble, monomeric protein with binding properties essentially identical to those of the intact receptor. The crystal structure of this protein has been determined in the presence and absence of various agonists and antagonists [Armstrong, N., Sun, Y., Chen, G. Q., and Gouaux, E. (1998) <i>Nature 395</i>, 913−917; Armstrong, N., and Gouaux, E. (2000) <i>Neuron 28</i>, 165−181]. The protein consists of two lobes, with the S1 segment composing the majority of lobe 1 and the S2 segment composing most of lobe 2. A domain closure upon ligand binding has been postulated, but details of intradomain motions have not been investigated. In this paper, the backbone motions of the ligand-binding core of GluR2 bound to glutamate were studied using <sup>15</sup>N longitudinal (<i>T</i><sub>1</sub>) and transverse (<i>T</i><sub>2</sub>) relaxation measurements as well as {<sup>1</sup>H}−<sup>15</sup>N nuclear Overhauser effects at 500 and 600 MHz. Residues in the agonist-binding pocket exhibited two main classes of motion. Those contacting the α-substituents of the ligand glutamate exhibited minimal internal motion, while those contacting the γ-constituents exhibited exchange dynamics, indicating two dynamically distinct portions of the binding pocket. Also, two residues in transdomain linkers between lobes 1 and 2 show exchange, lending new insight into the previously proposed domain closure hypothesis. Finally, concerted motion of helix F suggests a pathway for ligand dissociation without the necessity of domain reopening.