ReviewStructural insights into Cys-loop receptor function and ligand recognition
Graphical abstract
Introduction
Our expanding structural knowledge of membrane proteins, including ion channels, G-protein coupled receptors, transporters and pumps, progressively shifts the drug design process for these important drug targets from high-throughput screening toward a structure-based discovery approach. The treatment of neurological disorders could greatly benefit from rational drug development since many of these disorders could be better treated with more potent or efficacious drugs with fewer side effects. However, structure-based drug discovery approaches require a thorough insight into the structural features of target receptors, including the family of pentameric ligand-gated ion channels (pLGICs). This family consists of allosterically regulated membrane proteins located at synapses in the nervous system. They convert binding of a specific neurotransmitter released in the synaptic cleft into an ion flux over the postsynaptic membrane, which subsequently triggers excitatory or inhibitory postsynaptic potentials. Known vertebrate anion-selective pLGICs are the γ-amino butyric acid type A (GABAA), γ-amino butyric acid type C (GABAC) and glycine receptor, whereas the cation-selective pLGICs are the 5-HT3 serotonin and nicotinic acetylcholine (nACh) receptors. pLGICs serve as targets for a wide variety of frequently prescribed drugs, including smoking cessation aids, anxiolytics, anticonvulsants, muscle relaxants, hypnotics and anti-emetics. Dysfunction of pLGICs also plays an important role in several disorders of the central nervous system, including hyperekplexia [1], myasthenia gravis [2], epilepsy [3], irritable bowel syndrome (IBS) [4], Alzheimer's disease [5], schizophrenia [6] and Parkinson's disease [7].
The current understandings concerning structure, gating mechanism and ligand recognition of pLGICs (Fig. 1) are derived from electron microscopic imaging of the nACh receptor from the Torpedo marmorata (Tm) electric organ [8] and X-ray crystallographic studies of the extracellular domain (ECD) from the muscle α1 nACh receptor [9] and the water-soluble acetylcholine binding proteins (AChBP) [10]. Until 2005 it was assumed that this class of membrane proteins was uniquely expressed in multicellular eukaryotic organisms, but Tasneem et al. also identified several homologues in unicellular prokaryotic organisms (Fig. 2) [11]. In 2008 the first X-ray crystal structure of an integral prokaryotic pLGIC, derived from Erwinia chrysanthemi (ELIC) was determined [12]. This structure likely corresponds to a non-conducting conformation of the ion channel. About one year later a second crystal structure was determined for another prokaryotic homologue, derived from the cyanobacterium Gloeobacter violaceus (GLIC) [13], [14], which likely displays an open ion-conducting conformation. Both ELIC and GLIC form cation-selective ion channels, similar to the nACh receptor (nAChR) and the 5-HT3 receptor (5-HT3R). ELIC can be activated by primary amines such as GABA [15], [16], GLIC on the other hand responds to an increase in extracellular proton concentration [17]. In 2011 the first 3-dimensional structure of a eukaryotic, anion-selective pLGIC was determined. This glutamate-gated chloride (GluCl) channel is derived from Caenorhabditis elegans and reveals an open pore conformation, similar to GLIC [18].
Based upon this structural information, the overall architecture of pLGICs appears to be conserved and consists of five homologous or identical membrane-spanning subunits. Each of these subunits is composed of a N-terminal extracellular ligand binding domain, four transmembrane domains, an intracellular loop between the 3rd and 4th transmembrane domain and an extracellular C-terminus. They are also referred to as the family of Cys-loop receptors (CLRs) due to a highly conserved disulfide bond in one of the loops forming the interface between the extracellular ligand binding domain and the pore-forming transmembrane channel domain. This disulfide bond is not present in the prokaryotic homologues, but the architectural fold of this loop is similar to their eukaryotic homologues.
This review will outline the current structural knowledge and understandings concerning the molecular recognition of ligands, channel gating and ion permeation for this class of ion channels.
Section snippets
Ligand recognition in eukaryotic Cys-loop receptors
AChBPs are water-soluble homologues of the ligand binding domain (LBD) from eukaryotic pLGICs. They have been identified in several invertebrate organisms, including water snails, Lymnaea stagnalis (Ls-AChBP) [19], [20], Aplysia californica (Ac-AChBP) [21] and Bulinus truncatus (Bt-AChBP) [22] and the polychaete worm, Capitela teleta (Ct-AChBP) [23], where they modulate cholinergic transmission in the central nervous system [20]. Although the sequence identity among the different AChBPs is
Channel gating and ion permeation
Cys-loop receptor activation is the result of allosteric coupling between ligand binding and channel opening. The binding of an agonist to the ligand binding pocket in the extracellular domain has to be transmitted over a large distance (approximately 65 Å) to evoke opening of the ion channel pore, allowing ion permeation. Upon prolonged agonist binding, the channel however is transformed into a refractory, desensitized state that does not allow the permeation of ions. Upon dissociation of the
Conclusions and future challenges
This review discusses the functional features of pLGICs in the context of currently available structural insights for this group of ion channels, thereby providing a general outline of the current understanding of ligand recognition, channel gating and ion permeation for pLGICs.
High-resolution structures of AChBPs provide detailed insight into structural determinants of ligand recognition and conformational changes that possibly form initial triggers for channel opening. Crystal structures of
References (102)
- et al.
Myasthenia gravis
Lancet
(2001) - et al.
GABA(A) receptor epilepsy mutations
Biochem Pharmacol
(2004) - et al.
The role of nicotinic acetylcholine receptors in Alzheimer's disease
J Physiol Paris
(2006) Refined structure of the nicotinic acetylcholine receptor at 4A resolution
J Mol Biol
(2005)- et al.
Nicotine and carbamylcholine binding to nicotinic acetylcholine receptors as studied in AChBP crystal structures
Neuron
(2004) - et al.
Structural and ligand recognition characteristics of an acetylcholine-binding protein from Aplysia californica
J Biol Chem
(2004) - et al.
Allosteric mechanisms in normal and pathological nicotinic acetylcholine receptors
Curr Opin Neurobiol
(2001) - et al.
Nicotinic receptors in wonderland
Trends Biochem Sci
(2001) - et al.
Conserved tyrosines in the alpha subunit of the nicotinic acetylcholine receptor stabilize quaternary ammonium groups of agonists and curariform antagonists
J Biol Chem
(1994) - et al.
Mutational analysis of ligand-induced activation of the Torpedo acetylcholine receptor
J Biol Chem
(1992)
Functional significance of aromatic amino acids from three peptide loops of the alpha 7 neuronal nicotinic receptor site investigated by site-directed mutagenesis
FEBS Lett
Structural characterization of binding mode of smoking cessation drugs to nicotinic acetylcholine receptors through study of ligand complexes with acetylcholine-binding protein
J Biol Chem
Gating movement of acetylcholine receptor caught by plunge-freezing
J Mol Biol
Agonist-mediated conformational changes in acetylcholine-binding protein revealed by simulation and intrinsic tryptophan fluorescence
J Biol Chem
The F-loop of the GABA A receptor gamma2 subunit contributes to benzodiazepine modulation
J Biol Chem
Mutagenesis and molecular modeling reveal the importance of the 5-HT3 receptor F-loop
J Biol Chem
Ligand-specific conformational changes in the alpha1 glycine receptor ligand-binding domain
J Biol Chem
Structural rearrangements in loop F of the GABA receptor signal ligand binding, not channel activation
Biophys J
Conformational transitions underlying pore opening and desensitization in membrane-embedded Gloeobacter violaceus ligand-gated ion channel (GLIC)
J Biol Chem
Multi-site binding of a general anesthetic to the prokaryotic pentameric ligand-gated ion channel ELIC
J Biol Chem
Normal mode analysis suggests a quaternary twist model for the nicotinic receptor gating mechanism
Biophys J
An ion selectivity filter in the extracellular domain of Cys-loop receptors reveals determinants for ion conductance
J Biol Chem
In glycine and GABA(A) channels, different subunits contribute asymmetrically to channel conductance via residues in the extracellular domain
J Biol Chem
Genetic manipulation of ion channels: a new approach to structure and mechanism
Neuron
A prokaryotic perspective on pentameric ligand-gated ion channel structure
Curr Opin Struct Biol
A ring of uncharged constriction polar amino acids as a component of channel in the nicotinic acetylcholine receptor
FEBS Lett
Ligand-gated ion channels: mechanisms underlying ion selectivity
Prog Biophys Mol Biol
Mutational analysis of the charge selectivity filter of the alpha7 nicotinic acetylcholine receptor
Neuron
Desensitization mechanism in prokaryotic ligand-gated ion channel
J Biol Chem
Common determinants of single channel conductance within the large cytoplasmic loop of 5-hydroxytryptamine type 3 and alpha4beta2 nicotinic acetylcholine receptors
J Biol Chem
Structure of the M2 transmembrane segment of GLIC, a prokaryotic Cys loop receptor homologue from Gloeobacter violaceus, probed by substituted cysteine accessibility
J Biol Chem
Mutations in the alpha1 subunit of the inhibitory glycine receptor cause the dominant neurologic disorder, hyperekplexia
Nat Genet
Role of serotonin in the pathophysiology of the irritable bowel syndrome
Br J Pharmacol
The 5-HT 3 receptor as a therapeutic target
Expert Opin Ther Targets
Nicotinic receptors in aging and dementia
J Neurobiol
Crystal structure of the extracellular domain of nAChR alpha1 bound to alpha-bungarotoxin at 1.94 A resolution
Nat Neurosci
Crystal structure of an ACh-binding protein reveals the ligand-binding domain of nicotinic receptors
Nature
Identification of the prokaryotic ligand-gated ion channels and their implications for the mechanisms and origins of animal Cys-loop ion channels
Genome Biol
X-ray structure of a prokaryotic pentameric ligand-gated ion channel
Nature
Structure of a potentially open state of a proton-activated pentameric ligand-gated ion channel
Nature
X-ray structure of a pentameric ligand-gated ion channel in an apparently open conformation
Nature
Ligand activation of the prokaryotic pentameric ligand-gated ion channel ELIC
PLoS Biol
Pentameric ligand-gated ion channel ELIC is activated by GABA and modulated by benzodiazepines
Proc Natl Acad Sci U S A
A prokaryotic proton-gated ion channel from the nicotinic acetylcholine receptor family
Nature
Principles of activation and permeation in an anion-selective Cys-loop receptor
Nature
A glia-derived acetylcholine-binding protein that modulates synaptic transmission
Nature
Crystal structure of nicotinic acetylcholine receptor homolog AChBP in complex with an alpha-conotoxin PnIA variant
Nat Struct Mol Biol
Identification and functional characterization of a novel acetylcholine-binding protein from the marine annelid Capitella teleta
Biochemistry
Structures of Aplysia AChBP complexes with nicotinic agonists and antagonists reveal distinctive binding interfaces and conformations
EMBO J
Emerging structure of the nicotinic acetylcholine receptors
Nat Rev Neurosci
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