Review Lessons from more than 80 structures of the GluA2 ligand-binding domain in complex with agonists, antagonists and allosteric modulators Jacob Pøhlsgaard, Karla Frydenvang, Ulf Madsen, Jette Sandholm Kastrup * Department of Medicinal Chemistry, Faculty of Pharmaceutical Sciences, University of Copenhagen, Universitetsparken 2, DK-2100 Copenhagen, Denmark article info Article history: Received 15 April 2010 Received in revised form 22 July 2010 Accepted 7 August 2010 Keywords: GluA2 Ligand-binding domain Agonist Antagonist Positive allosteric modulator Crystal structure abstract Ionotropic glutamate receptors (iGluRs) constitute a family of ligand-gated ion channels that are essential for mediating fast synaptic transmission in the central nervous system. These receptors play an important role for the development and function of the nervous system, and are essential in learning and memory. However, iGluRs are also implicated in or have causal roles for several brain disorders, e.g. epilepsy, Alzheimer’s disease, Parkinson’s disease and schizophrenia. Their involvement in neurological diseases has stimulated widespread interest in their structure and function. Since the first publication in 1998 of the structure of a recombinant soluble protein comprising the ligand-binding domain of GluA2 extensive studies have afforded numerous crystal structures of wildtype and mutant proteins including different ligands. The structural information obtained combined with functional data have led to models for receptor activation and desensitization by agonists, inhibition by antagonists and block of desensi- tization by positive allosteric modulators. Furthermore, the structural and functional studies have formed a powerful platform for the design of new selective compounds. Ó 2010 Elsevier Ltd. All rights reserved. 1. Introduction to the ionotropic glutamate receptors Glutamate (Scheme 1) is the principal excitatory neurotrans- mitter within the mammalian central nervous system (CNS). There are two major types of glutamate receptors, the metabotropic and ionotropic glutamate receptors (iGluRs) (Jingami et al., 2003; Kew and Kemp, 2005). The iGluRs constitute a family of ligand-gated ion channels that are essential for mediating fast synaptic trans- mission in the CNS. These receptors play an important role for the development and function of the nervous system, and are essential in learning and memory (Dingledine et al., 1999; Meldrum, 2000). However, iGluRs are also implicated in or have causal roles for several brain disorders, e.g. epilepsy, stroke, Alzheimers disease, Parkinsons disease, schizophrenia and neuropathic pain (Madsen et al., 2005; Foster and Kemp, 2006; Bowie, 2008). Their involve- ment in neurological diseases has stimulated widespread interest in their structure and function. The iGluRs have been divided into three subtypes according to their sequence identities and sensitivity to the agonists 3-amino- 3-(3-hydroxy-5-methyl-4-isoxazolyl)propionic acid (AMPA), kai- nate or N-methyl-D-aspartate (NMDA) (Collingridge et al., 2009; Lodge, 2009). The AMPA subtype comprises four different subunits (GluA1-GluA4), and these iGluRs are the main receptors responsible for fast excitatory signaling in the mammalian CNS. Until recently, the structural investigations on AMPA receptors were limited to studies of soluble constructs of individual domains, e.g. the N-terminal domain (Jin et al., 2009) and the ligand-binding domain (LBD) (GluA2: Armstrong and Gouaux, 2000; GluA3: Ahmed et al., 2009a and GluA4: Kasper et al., 2008). However, the crystal structure of the rat full-length homotetrameric GluA2 receptor was determined to 3.6 Å resolution in complex with the competitive antagonist ZK200775 (Sobolevsky et al., 2009) (pdb- code 3KG2). The receptor shows an overall axis of two-fold symmetry but with the ion channel domain displaying four-fold symmetry. Both the extracellular N-terminal domains and the LBDs are organized in layered pairs of local dimers. Interestingly, a cross- over of extracellular domains from one subunit to the other is observed (Fig. 1A). The recombinant soluble protein of the LBD of GluA2 (Chen and Gouaux, 1997; Chen et al., 1998; Armstrong et al., 1998 (pdb-codes 1GR2 and 1FTK); Armstrong and Gouaux, 2000) has been extensively studied since the first publication in 1998 of the structure, and crystal structures combined with functional data have led to models for receptor activation and desensitization by agonists and inhibition by antagonists (Armstrong and Gouaux, 2000; Sun et al., 2002; Jin et al., 2003, 2005; Armstrong et al., 2006). The desensitized state is a state in which the agonist remains bound but the ion channel is closed. * Corresponding author. Tel.: þ45 3533 6486; fax: þ45 3533 6001. E-mail address: jsk@farma.ku.dk (J.S. Kastrup). Contents lists available at ScienceDirect Neuropharmacology journal homepage: www.elsevier.com/locate/neuropharm 0028-3908/$ e see front matter Ó 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.neuropharm.2010.08.004 Neuropharmacology 60 (2011) 135e150