Structure and gating of tetrameric glutamate receptors

Abstract

Ionotropic glutamate receptors (iGluRs) are ligand-gated ion channels that open their ion-conducting pores in response to the binding of agonist glutamate. In recent years, significant progress has been achieved in studies of iGluRs by determining numerous structures of isolated water-soluble ligand-binding and amino-terminal domains, as well as solving the first crystal structure of the full-length AMPA receptor in the closed, antagonist-bound state. These structural data combined with electrophysiological and fluorescence recordings, biochemical experiments, mutagenesis and molecular dynamics simulations have greatly improved our understanding of iGluR assembly, activation and desensitization processes. This article reviews the recent structural and functional advances in the iGluR field and summarizes them in a simplified model of full-length iGluR gating.

Figure 1. Structure of iGluR

A, topology of iGluR subunit. B, structure of AMPA subtype rat GluA2 receptor in the closed antagonist bound state (3KG2). The four subunits (A to D) are coloured differently. The antagonist ZK 200775 molecules are shown as space-filling models. Strong and weak interfaces are shown as large and small ovals, respectively. C, model of the assembly of four three-compartmental sausages with strong interactions holding together (1) all four bottom compartments, (2) left and right pairs of the top compartments and (3) front and back pairs of the middle compartments. D, LBD–TMD linkers – the iGluR gating transmission domain – include S1–M1, M3–S2 and S2–M4, of which S1–M1 and S2–M4 are shown transparent. The M3–S2 linkers, the central element of the gating machinery, have different conformations and secondary structures for the two diagonal pairs of subunits, A/C and B/D.

Figure 2. Gating at the level of LBD

A, whole-cell current recorded from HEK 293 cell expressing rat GluA2i. The current (courtesy of Dr Yelshansky, Columbia University) was elicited by a 500 ms application of glutamate (3 mm; filled bar) at a holding potential of −60 mV. B–D, back-to-back LBD dimers from the closed state structure of GluA2 (3KG2) bound to ZK 200775 (B) and from structures of the glutamate-bound isolated S1S2 in the presumed open (1FTJ; C) and desensitized (S729C mutant, 2I3W; D) states. The α-helical elements of secondary structure are labelled in B. Shown are distances between Cα's (spheres) of P632 and G739. Putative gating-associated movements of D1 and D2 are illustrated by red arrows.

Figure 3. Ion channel

A, GluA2 ion channel (3KG2) viewed parallel to membrane. The subunits are coloured similarly to Fig. 1B. The transmembrane segments M1 to M4 and the calf helix pre-M1 are labelled. B, superposition of the GluA2 channel (3KG2, cyan) and KcsA (1BL8, orange). C, GluA2 channel viewed from the extracellular side of membrane. The diagonal pairs of subunits, A/C and B/D, are shown in light green and purple, respectively. Molecular surface representation emphasizes that the channel is in the closed conformation. D, Shaker K⁺ channel in the open state (2A79) viewed from the intracellular side of membrane. Colouring of subunits is similar to the GluA2 channel in C. Comparing the structures of the closed GluA2 channel in C and the open Shaker channel in D, one can imagine how transmembrane helices of iGluR can bend and splay away from the central axis of the channel, mimicking the iris-like opening of K⁺ channels.

Figure 4. Structural model of iGluR gating

Shown is the full-length iGluR in the presumed closed, open and desensitized states. The closed state is represented by the GluA2 crystal structure (3KG2), while open and desensitized states are putative conformations constructed based on a set of assumptions (see main text). Simulation of opening movements of the ion channel segments homologous to the corresponding segments of the Shaker K⁺ channel (2A79) and docking of the ion channel, LBDs and ATDs relative to each other in the open and desensitized states were carried out using the program Superpose (CCP4 program suite). The diagonal pairs of subunits, A/C and B/D, are shown in light green and purple, respectively. Vertical and equatorial arrow symbols indicate shortening/elongation and twisting/untwisting changes in the structure, respectively. The arrow thickness corresponds to the motion strength: the thicker the arrow the stronger motion. Note, twisting and shortening of the structure occur during opening, while untwisting and elongation during desensitization (see Supplemental Movie 3). Transitions between the closed, open and desensitized states that are shown as unidirectional can potentially be reversed.