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Fig. 1. Predicted topology of a GABAA receptor subunit. The cysteine-disulfide bridge in the N-terminus is indicated by a black bar. Transmembrane domains are shown as open boxes labelled TM1-4.

of the TM2 domains and includes part of the TM1-TM2 loop. Mutating these amino acids in the a 7 subunit of homomeric nicotinic receptors produced acetylcholine-gated anion channels (Galzi et al., 1992). The converse can also be done: mutation of five amino acids in the TM1-TM2 loop of the GABAa receptor p3 subunit to the corresponding amino acids of the a7 nicotinic acetylcholine subunit produces cation-selective GABAA receptors (Jensen et al., 2002). Similar mutations in the a2 or g2 subunits did not change ion selectivity. Thus the p subunits predominantly determine the ion selectivity of the GABAA receptor (Jensen et al., 2002).

In the 5-HT3 and nicotinic receptors, residues in the TM3-TM4 loop region influence single channel conductance (Peters et al., 2005; Hales et al., 2006). The TM3-TM4 loop, which may be relatively unstructured in the nAChR (Kukhtina et al., 2006), contributes key sites for attaching anchor and regulatory proteins involved in locating the receptor at synapses and in governing the activity of GABAA receptors (Kittler and Moss, 2003) (see Sections "Synaptic GABAA receptors: apg subunit combinations and anchoring role of the g2 subunit and gephyrin'' and "Regulation of GABAA receptor function by neuromodulators: the role of kinases and phosphatases"). But the TM4 region of the g2 subunit is necessary and sufficient to confer a synaptic localization on the receptor (Alldred et al., 2005).

The atomic structure of a GABAA receptor subunit complex has not so far been solved directly. Instead, realistic models have used the empirically determined structural coordinates of the muscle nicotinic acetylcholine receptor from the electric organ of the Torpedo ray fish and a related snail acetylcholine receptor binding protein (AChBP) (Brejc et al., 2001; Cromer et al., 2002; Ernst et al., 2003; Unwin, 2003, 2005). The AChBP shows sequence similarity with the N-terminus of the nAChR at regions that build the agonist-binding sites; the AChBP contains a Cys-loop but lacks the transmembrane domains. It assembles as soluble homopentamers (Brejc et al., 2001). The crystal structure of the AChBP, with bound ligand, provided a template for comparative modelling of the N-terminal extracellular domain of GABAA

receptors (Ernst et al., 2003), whereas Unwin's most recent structure of the Torpedo nAChR at 4 A resolution obtained by cryo-EM, and incorporating insights from the AChBP, has given a full-scale atomic model (Protein Data Bank Code 2BG9). According to Xiu and colleagues, 2BG9 represents a substantial advance for the field, and all modern attempts to obtain molecular scale information on the structure and function of Cys-loop receptors must consider this as a starting point (Xiu et al., 2005; Unwin, 2005). 2BG9 provides us with a view of how the entire GABAA receptor must look, including the transmembrane and large cytoplasmic loops (Unwin, 2003, 2005).

Before 2BG9, modellers used family conservation patterns and fold predictions to estimate that 60-75% of the amino acid residues of the GABAA receptor subunits have structural equivalents in the AChBP template (Ernst et al., 2003). The accuracy of the GABAA receptor model will be limited in regions where alignment is unclear (e.g., due to low sequence identity) or in regions where the AChBP differs from other family members due to its soluble, non-membrane-bound nature (Ernst et al., 2003). A model of the extracellular domain of a pentameric GABAA receptor consisting of two a, two p and one g2 subunit is shown in Fig. 2. In this model the amino acids known to contribute to ligand-binding sites and interfaces are correctly positioned and the interface-forming segments and the solvent accessibility of individual residues correlate well with experimental data (Ernst et al.,

2003). Six "loops" (loop A, B, C for the plus side and D, E, F for the minus side) at the interface between neighbouring subunits form the ligand-binding sites (Sigel and Buhr, 1997; Olsen et al.,

2004). The binding pocket for GABA forms at the interface between the a and the p subunit (Figs. 2a, c, d), the binding pocket for benzodiazepines lies at the interface of the a and the g subunit (Figs. 2a, b). The predicted space for agonist binding is formed by loops A, B, C, D and E (blue volume in Fig. 2d) and correlates with experimental data from photo-labeling of a1F64 by [3H]muscimol and substituted cysteine accessibility mapping (Ernst et al., 2003; Olsen et al., 2004). Amino acid residues on loops A, B, C, D and E at the interface of the a and the g subunit influence binding, potency and efficacy of

Binding Potency

Fig. 2. Model of the extracellular domains of a pentameric GABAa receptor consisting of two a, two b and one g2 subunit. (a) View from the extracellular space. GABA binds to the interface between the a and the b subunit, benzodiazepines bind to the interface between the a and the g2 subunit. (b) Predicted benzodiazepine-binding pocket between the a and the g2 subunit, viewed from the side. The binding site loops are labelled A to G. (c) and (d) The a and b subunit viewed from the side. Loops A, B, C, D and E form the predicted GABA-binding pocket (blue volume in (d)). The volume shown in green might be used in antagonist-bound states. (Adapted from Ernst et al., 2003 used with permission.)

Fig. 2. Model of the extracellular domains of a pentameric GABAa receptor consisting of two a, two b and one g2 subunit. (a) View from the extracellular space. GABA binds to the interface between the a and the b subunit, benzodiazepines bind to the interface between the a and the g2 subunit. (b) Predicted benzodiazepine-binding pocket between the a and the g2 subunit, viewed from the side. The binding site loops are labelled A to G. (c) and (d) The a and b subunit viewed from the side. Loops A, B, C, D and E form the predicted GABA-binding pocket (blue volume in (d)). The volume shown in green might be used in antagonist-bound states. (Adapted from Ernst et al., 2003 used with permission.)

benzodiazepines (see Section "GABAA receptors: allosteric modulation by benzodiazepines and related ligands'') (Fig. 2b) (Ernst et al., 2003). The predicted benzodiazepine pocket is larger than the GABA pocket. It communicates with the Cys-loop of the a subunit and extends down to the membrane-near part, which possibly contains side chains from the linker between transmembrane region 2 and 3 of the a subunit (Ernst et al., 2003).

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