Iehab N. Talukder

2nd Year Graduate Student

Department: Neurobiology & Behavior

Graduate Program: Neuroscience

Advisor: Lonnie Wollmuth


Abstract:

Title: Coupling of ligand binding to ion channel gating in the NMDA receptor.

Iehab Talukder, Michael Prodromou, Priya Borker, Lonnie P. Wollmuth.

The N-methyl-D-aspartate (NMDA) receptor mediates a critical component of glutamate-induced fast synaptic transmission in the mammalian central nervous system. Moreover, physiologic activity of the NMDA receptor is implicated in the induction of synaptic plasticity- the cellular parallel of learning and memory. Under normal circumstances, synapses maintain a precise level of NMDA receptor activity. Disruption of the balance between over- and under-activity of the NMDA receptor is evident in several disease states, including stroke, neurodegenerative disease and mental disorders. Maintenance of this balance has also proved to be elusive by numerous attempts of drug design that seeks to adjust pathologic levels of NMDA receptor activity.

The NMDA receptor is a heterotetrameric ion channel most often composed of two NR1 and two NR2 subunits. Each NMDA receptor subunit harbors an N terminal zinc-binding modulatory domain and a ligand-binding domain, both located extracellularly. The transmembrane ion channel is composed of three membrane-spanning segments M1, M3 and M4, as well as a reentrant M2 pore loop. M2 and M3 are the major pore-lining segments. The ligand-binding domain (LBD) comprises lobes S1 (N terminal to M1) and S2 (between M3 and M4). The three 12-20 amino acid long segments attaching the ligand-binding domain to the ion channel are the linker regions S1-M1, M3-S2 and S2-M4. Additionally, there is a C terminal domain involved in intracellular signal transduction and trafficking.

We have used the substituted cysteine accessibility method (SCAM) on NR1-NR2C receptors to investigate the relative dynamics of the linker regions in the agonist bound and unbound states. Analysis of the discrete changes in whole-cell currents carried by the modified receptors have led us to infer about the relative movements of the linker regions during gating, as well as possible mechanisms of these movements. We show that, in agreement with current knowledge of the structural dynamics of the LBD and transmembrane ion channel, the M3-S2 linker undergoes the most extensive motion during receptor gating. We have also identified a putative helical interface in the S2-M4 linker, which we propose is the interacting surface for the M3-S2 linker. This interaction between the relatively immobile S2-M4 and the dynamic M3-S2 linkers modulates the gating properties of the NMDA receptor. We hope to exploit this interaction to stabilize the linker regions as we try to crystallize them.