nomenclature, the pentameric ligand-gated ion channels (pLGICs; a better term because prokaryotic
homologues lack the characteristic cystine bridge of their eukaryotic counterparts). Other members of
the family include the vertebrate GABA A , glycine, and excitatory serotonin (5-HT 3 ) receptors, as well
as an invertebrate glutamate-gated chloride channel (GluCl). The ACh and 5-HT 3 receptors gate
cationic channels, GluCl, and the GABA A and glycine receptors anionic ones. Orthologues have
been discovered in prokaryotes, the bacterial channels GLIC and ELIC, which have played a critical
role in crystallographic studies of the pLGIC superfamily (for reviews, see e.g. Corringer et al. 2012;
Sine 2012; Nys et al. 2013; Unwin 2013; Taly et al. 2014; Cecchini and Changeux 2015; Changeux
et al. 2015; Nemecz et al. 2016).
The overall organization common to all pLGICs was first established by cryo-EM studies,
carried out by Nigel Unwin and his colleagues, of the nAChR from the electric ray, Torpedo
marmorata. Torpedo electrocytes, which are evolutionarily derived from muscle fibers, produce
massive amounts of receptors, which are densely packed in the postsynaptic membrane. The receptor
present in purified postsynaptic membrane fragments can be prodded into organizing into tubular 2D
crystals, in which it adopts a variety of helical arrangements suitable for crystallographic analysis. The
study of cryo-EM images of the tubes has shown that the five subunits are arranged pseudosymmetrically around an axis normal to the membrane, along which runs the cation-selective channel (Brisson
and Unwin 1985; see Unwin 2013, and references therein). The walls of the channel are formed by TM
helices contributed by each subunit (Figs. 1.26, right, and 1.27, left).
The best electron density maps obtained by cryo-EM studies of Torpedo helical tubes are limited
to ~4-Å resolution (Unwin 2005, 2013; Unwin and Fujiyoshi 2012). The nAChR receptor itself has
resisted protracted attempts at 3D crystallization, but a host of high-resolution data have been obtained
by (i) X-ray studies of a non-TM ACh-binding protein extracted from the central nervous system of the
mollusc Lymnaea stagnalis, which is homologous to the extracellular region of the nAChR (Brejc et al.
2001; Smit et al. 2001); (ii) X-ray studies of the bacterial pLGICs ELIC, from Erwinia chrysanthemi
(Hilf and Dutzler 2008; Spurny et al. 2012), and GLIC, from Gloeobacter violaceus (Bocquet et al.
2009; Hilf and Dutzler 2009; Nury et al. 2011; Prévost et al. 2012; Sauguet et al. 2013, 2014); (iii)
X-ray studies of the GluCl channel from the invertebrate Caenorhabditis elegans (Hibbs and Gouaux
2011; Althoff et al. 2014), of the human GABA A receptor (Miller and Aricescu 2014) and of the mouse
5-HT 3 receptor (Hassaine et al. 2014); (iv) single-particle cryo-EM studies of the glycine receptor from
zebrafish (Du et al. 2015); and (v) the X-ray structure of a central nervous system ACh receptor, the
α4β2 nicotinic receptor (Morales-Perez et al. 2016). Taken together, these data provide a detailed view
of how binding of the ligand controls the opening and closing of the channel. Whereas some details
may differ from one protein to the other, the general picture that emerges from this ensemble of data
gathered on disparate systems is generally very consistent.
In short, the binding of agonists induces a reorganization of the extramembrane region, which
has been described as resulting from the composition of a radial movement (the pentamer “blooms” in
the closed vs. the open state of the receptor; Fig. 1.28A) and a twisting movement (Fig. 1.28B). This
entails a rearrangement of the interface between the extramembrane region and the TM helix bundle,
pushing slightly outward the outermost part of the pore-lining M2 helices, which opens the pore.
In Fig. 1.29 is shown a model (Calimet et al. 2013; Taly et al. 2014) of the mechanism of signal
transduction between the extramembrane and TM regions of pLGICs, based on MD simulations using
the structures of the prokaryotic channels GLIC and ELIC and the eukaryotic channel GluCl. Residues
belonging to the extramembrane region act on the M2-M3 loop of the TM helix bundle to control the
position of the outermost part of helix M2, thereby opening and closing the pore. According to
experimental data, the displacement of M2 is small and has a limited impact on the positions of the
other helices, entailing only small movements at the interface with the membrane in GluCl and GlyR,
and essentially none in GLIC.
1.6 Dynamics of Transmembrane Regions and the Function of Membrane Proteins
39
homologues lack the characteristic cystine bridge of their eukaryotic counterparts). Other members of
the family include the vertebrate GABA A , glycine, and excitatory serotonin (5-HT 3 ) receptors, as well
as an invertebrate glutamate-gated chloride channel (GluCl). The ACh and 5-HT 3 receptors gate
cationic channels, GluCl, and the GABA A and glycine receptors anionic ones. Orthologues have
been discovered in prokaryotes, the bacterial channels GLIC and ELIC, which have played a critical
role in crystallographic studies of the pLGIC superfamily (for reviews, see e.g. Corringer et al. 2012;
Sine 2012; Nys et al. 2013; Unwin 2013; Taly et al. 2014; Cecchini and Changeux 2015; Changeux
et al. 2015; Nemecz et al. 2016).
The overall organization common to all pLGICs was first established by cryo-EM studies,
carried out by Nigel Unwin and his colleagues, of the nAChR from the electric ray, Torpedo
marmorata. Torpedo electrocytes, which are evolutionarily derived from muscle fibers, produce
massive amounts of receptors, which are densely packed in the postsynaptic membrane. The receptor
present in purified postsynaptic membrane fragments can be prodded into organizing into tubular 2D
crystals, in which it adopts a variety of helical arrangements suitable for crystallographic analysis. The
study of cryo-EM images of the tubes has shown that the five subunits are arranged pseudosymmetrically around an axis normal to the membrane, along which runs the cation-selective channel (Brisson
and Unwin 1985; see Unwin 2013, and references therein). The walls of the channel are formed by TM
helices contributed by each subunit (Figs. 1.26, right, and 1.27, left).
The best electron density maps obtained by cryo-EM studies of Torpedo helical tubes are limited
to ~4-Å resolution (Unwin 2005, 2013; Unwin and Fujiyoshi 2012). The nAChR receptor itself has
resisted protracted attempts at 3D crystallization, but a host of high-resolution data have been obtained
by (i) X-ray studies of a non-TM ACh-binding protein extracted from the central nervous system of the
mollusc Lymnaea stagnalis, which is homologous to the extracellular region of the nAChR (Brejc et al.
2001; Smit et al. 2001); (ii) X-ray studies of the bacterial pLGICs ELIC, from Erwinia chrysanthemi
(Hilf and Dutzler 2008; Spurny et al. 2012), and GLIC, from Gloeobacter violaceus (Bocquet et al.
2009; Hilf and Dutzler 2009; Nury et al. 2011; Prévost et al. 2012; Sauguet et al. 2013, 2014); (iii)
X-ray studies of the GluCl channel from the invertebrate Caenorhabditis elegans (Hibbs and Gouaux
2011; Althoff et al. 2014), of the human GABA A receptor (Miller and Aricescu 2014) and of the mouse
5-HT 3 receptor (Hassaine et al. 2014); (iv) single-particle cryo-EM studies of the glycine receptor from
zebrafish (Du et al. 2015); and (v) the X-ray structure of a central nervous system ACh receptor, the
α4β2 nicotinic receptor (Morales-Perez et al. 2016). Taken together, these data provide a detailed view
of how binding of the ligand controls the opening and closing of the channel. Whereas some details
may differ from one protein to the other, the general picture that emerges from this ensemble of data
gathered on disparate systems is generally very consistent.
In short, the binding of agonists induces a reorganization of the extramembrane region, which
has been described as resulting from the composition of a radial movement (the pentamer “blooms” in
the closed vs. the open state of the receptor; Fig. 1.28A) and a twisting movement (Fig. 1.28B). This
entails a rearrangement of the interface between the extramembrane region and the TM helix bundle,
pushing slightly outward the outermost part of the pore-lining M2 helices, which opens the pore.
In Fig. 1.29 is shown a model (Calimet et al. 2013; Taly et al. 2014) of the mechanism of signal
transduction between the extramembrane and TM regions of pLGICs, based on MD simulations using
the structures of the prokaryotic channels GLIC and ELIC and the eukaryotic channel GluCl. Residues
belonging to the extramembrane region act on the M2-M3 loop of the TM helix bundle to control the
position of the outermost part of helix M2, thereby opening and closing the pore. According to
experimental data, the displacement of M2 is small and has a limited impact on the positions of the
other helices, entailing only small movements at the interface with the membrane in GluCl and GlyR,
and essentially none in GLIC.
1.6 Dynamics of Transmembrane Regions and the Function of Membrane Proteins
39