See also Receptor.

Cys-loop receptors

Cationic cys-loop receptors

Serotonin type-3 receptor (5-HT3-R)

• 5-hydroxytryptamine receptor#Structural highlights/Specific Function of 5-HT3

The 5-HT3 receptor is a pentameric cation-selective ion channel and plays a role in neuronal excitation to release neurotransmitters from the postsynaptic neuron. Opening of the cation channel causes an influx of sodium and calcium through the receptor pore leading to a membrane depolarization. Five receptor subunits, A to E, have been found in humans but only subunits A and B have been found in rodents. When experimentally expressed in a host, the 5-HT3 receptor is comprised of either A or AB subunits which can result in a homopentameric receptor or a heteropentameric receptor respectively. The A and B subunits are found throughout the brain in areas such as the hippocampus and amygdala. 5-HT3 is a transmembrane channel that is stimulated to open state by the interaction of the receptor with serotonin in the extracellular space. The binding site is comprised of 6 loops from 2 adjacent subunits in the extracellular N-terminal domain. Loops A, B and C form the principal subunit and contain the important side chains N128, W183 and Y234. Loops D, E and F form the complementary subunit of the binding site and contain the important side chains W90, Y143 and W195. The transmembrane region is comprised of multiple alpha helical structures and mediates ion flow and ion specificity.

• The extracellular subunit interface of the 5-HT3 receptors: a computational alanine scanning mutagenesis study^[1]

The serotonin type-3 receptor is a cation selective transmembrane protein channel that belongs to the Cys–loop Ligand-Gated Ion Channel (LGIC) superfamily, which also includes receptors for nicotinic acetylcholine (nAChR, PDB code 2bg9), γ-aminobutyric acid and glycine. 5-HT3-R is involved in signal transmission in the central and peripheral nervous system and its malfunctioning leads to neurodegenerative and psychiatric diseases, therefore it is an important target for drug design research. A few drugs active against 5-HT3-R are already on the market, such as, for example, palonosetron (http://en.wikipedia.org/wiki/Palonosetron) and granisetron (http://en.wikipedia.org/wiki/Granisetron). The 5-HT3R is made of 5 monomers assembled in a pseudo-symmetric pentameric shape to form an ion channel permeable to small ions (Na+, K+); each subunit contains 3 domains: an intracellular portion, a transmembrane domain and an extracellular region (shown on the example of nAChR, 2bg9). To date, 5 different 5-HT3-R subunits have been identified, the 5-HT3 A, B, C, D and E; however, only subunits A and B have been extensively characterized experimentally. The ligand binding site of nAChR is located at the extracellular region, at the interface between 2 monomers (α-γ and α-δ; 2 identical α monomers, chains A and D, are colored in same color - lavender), called the principal and the complementary subunits. The 3D structure of 5-HT3-R has not been experimentally solved yet; however, it has been obtained computationally by means of homology modelling techniques. (http://salilab.org/modeller/) Thus, the extracellular region of the 5HT3 subunits A and B are modelled by homology with the 3D structure of the nAChR subunit A (2bg9-A) and are used to assemble receptor structures as pseudo-symmetric pentamers made either of five identical subunits A (homomeric 5-HT3A-R) or of both subunits A and B (heteromeric 5-HT3A/B-R in the BBABA arrangement) in a still debated arrangement.^[2] Subunits A and B are colored in magenta and red, respectively. A complete characterization of the extracellular moiety of the dimer interface of the 5-HT3-R (AA dimer is shown, principal subunit is colored in cyan and complementary is in blue, is obtained by the Computational Alanine Scanning Mutagenesis (CASM) approach ^[3], which simulates the substitution, one by one, of all the amino acid residues at the subunit-subunit interfaces with an Ala, thus to assess the interface binding contribution of single residue side-chains. The most relevant residues for interface stabilization are classified as “hot spots” that stabilize the interface by more than 4 kcal/mol and “warm spots” that contribute to interface stabilization by more than 2 kcal/mol. Click here to see also the interface of complementary subunit. Interface residues are shown in spacefill representation, hot spot residues are colored in red and warm spots residues are are in orange. From this analysis the important aromatic cluster located at the interface core and formed by residues W178 (principal subunit), Y68, Y83, W85 and Y148 (complementary subunit) is highlighted.^[4] In addition, 2 important groups of interface residues are probably involved in the coupling of agonist and antagonist binding to channel activation/inactivation: W116-H180-L179-W178-E124-F125 (principal subunit) and Y136-Y138-Y148-W85-(P150) (complementary subunit), where W178 and Y148 appear to be critical residues for the binding/activation mechanism. Finally, the comparison of the AA interface with the BB interface (principal subunit of AA is colored in cyan, principal subunit BB is colored in darkmagenta, complementary subunit AA is in blue and complementary subunit BB is in magenta) shows differences which could explain the reasons why the homopentamer 5-HT3B-R, if expressed, is not functional.^[5]

• 5-hydroxytryptamine receptor#5-HT3 receptor antagonists

5-HT3A receptor

The 5-HT3 receptor is bullet-shaped and consists of 5 subunits (A-E) that form an oligomer. In the center of this pentamer of is a ligand-gated ion channel full of water, which the 5 subunits enclose pseudo-symmetrically. Each subunit of the 5-HT3 receptor consists of 3 regions; the extracellular region, the transmembrane region, and the intracellular region. The extracellular region is relatively large compared to the other 2 regions, and contains a short C-terminus and a larger N-terminus. The N-terminus of the extracellular region is where the ligand binding occurs, and therefore deals with the agonists and antagonists. These binding sites are located between 2 bordering subunits, assembled from 3 α-helices of 1 subunit and 3 β-strands from the other subunit. Such connection creates a binding pocket with a small number of residues from each subunit pointed into the binding pocket, as opposed to the large number of residues that are pointing away from the binding pocket. This binding pocket shrinks around agonists, encapsulating them, and widens around antagonists, repulsing them. The transmembrane region is within the C-terminus region, and contains 4 α-helical domains (M1-M4) that stretch the length of this inner, transmembrane area. These 4 α-helical domains conduct the channel openings via ion selectivity, depending on both charge and size. M2, the porous domain, contains rings of charged amino acids at both its start and its end, accounting for M2 main contribution to ion selectivity. The M3 and M4 α-helices create a large loop with one another, thus assembling the intracellular region.

Nicotinic acetylcholine receptors

• Nicotinic Acetylcholine Receptors in general

The receptor is a transmembrane pentameric glycoprotein. It cylindrical in appearance by electron microscopy approximately 16nm in length and 8nm in diameter. The main ion channel is composed of a water pore that runs through the entire length of the protein. If viewed from the synaptic cleft, the protein will look like a pseudo-symmetrical rosette shown in the picture below composed of 10 different alpha and 4 different beta subunits.

• Side view.
• View from extracellular side.
• View from cytoplasmic side.

• Alpha-bungarotoxin is a nicotinic cholinergic antagonist that is found within the venom of Bungarus multicinctus, a South-asian snake.
• Binding site of AChR
• Acetylcholine Receptor and its Reaction to Cobra Venom

When cobra venom is introduced into the body is moves along the bloodstream to a diaphragm muscle. It works as a postsynaptic neurotoxin binding to the receptor as an extracellular ligand by interacting with OH group leaving the acetylcholine channel open which releases ions used in creating an action potential. There must be 5 molecules of cobra toxin (red) to block the receptor (blue) as each molecule binds with an individual alpha chain on the acetylcholine receptor. The 2nd image depicts an individual toxin binding with one chain on the receptor, both in the same color. Cobra Venom Interaction with Acetylcholine Receptor. This representation shows each molecule of the Cobra toxin binding to one chain of the receptor.

Anionic cys-loop receptors

GABA_A receptors

See GABAA receptor

Ionotropic Glutamate Receptors

AMPA glutamate receptor

• AMPA glutamate receptor by University of Massachusetts Amherst Chemistry-Biology Interface Program at UMass Amherst and on display at the Molecular Playground.

Full view of the glutamate receptor shows the overall structure (N-terminal, ligand-binding and transmembrane domains) in ribbon and spacefilling models. N-terminal domain is a part of the extracellular domain. This domain is implicated in receptor assembly, trafficking, and localization.

• Transmembrane Domain.
• Transmembrane Domain, other representaion. This domain widens in response to glutamate binding allowing for positive ions to pass through the post-synaptic membrane.
• Receptor antagonist 2K200225 binding site.
• Glutamate binding site.
• Glutamate receptor (GluA2)

The homomeric rat GluA2 receptor has 4 subunits arranged in a 'Y'-shape with the 'top' being about 3 times the width of the 'bottom'. This structure is a functional homotetramer of the AMPA-subtype; native ionotropic glutamate receptors are almost exclusively heterotetramers. Fancy, high-quality cartoons on/off.

Domains

The subunits themselves are modular ^[6]and the major domains are found in layers in the tetrameric structure.

• The 'top' layer is composed of the amino-terminal domain (ATD)

This extracellular domain is glycosylated.

• The ligand-binding domain (LBD) participates directly in agonist/competitive antagonist binding, affects activation gating, and is the portion that forms the 'middle' layer.

The competitive antagonist ZK200775 is bound to the LBD in the structure.

The ZK200775, a phosphonate quinoxalinedione AMPA antagonist^[7], was studied as a treatment for stroke because it had demonstrated neuroprotective efficacy in experimental models of stroke and tolerability in healthy volunteers; however, in a multicenter, double-blind, randomized, placebo-controlled phase II trial, it was found to have significant sedative effects in patients with acute stroke which precludes its further development as a neuroprotective agent^[8].

• The transmembrane domain (TMD) is the portion that forms the membrane-spanning on the 'bottom' of the solved structure.

To help give a better idea of how the glutamate receptor is oriented on the cell surface in the membrane lipid bilayer, a slab representative of hydrophobic core of the lipid bilayer as calculated by the Orientations of Proteins in Membranes database (University of Michigan, USA) is shown with the red patch of spheres indicating the boundary of the hydrophobic core closest to the outside of the cell and the dark blue patch of spheres indicating the boundary closest to the inside of the cell.

• The carboxy-terminal domain that plays a role in both receptor localization and regulation is not seen in the structure but would be below the transmembrane domain as it is cytoplasmic.

Domain swapping between the subunits and symmetry mismatch between the domains

• Unanticipated is the domain swapping and crossover that occurs between the subunits interactions. In order to discuss the remarkable swapping, it is best to designate each subunit with a letter:
      A       B       C        D

• Considering each chain, there is crossover as the pairs of subunits seen in the ATD are swapped in the LBD.

In the ATD domain -

• Portions of the A and B subunits pair up.
• And the Portions of the C and D subunits form a pair.
• While that is going on, in the ATD there is also inter-pair interactions mediated between subunits B and D. Note this view really highlights the two-fold symmetry between the A-B and C-D pairs at the level of the ATD.

In the LBD domain -

• Whereas in the ATD domain A and B paired up, portions of the A and D subunits pair up in the LBD.
• And the Portions of the B and C subunits form a pair.
• While that is going on, in the LBD there is also extensive inter-pair interactions mediated between subunits A and C. Note this view highlights the two-fold symmetry between the A-D and B-C pairs at the level of the LBD. Looking from the side helps in seeing the inter-pair interactions between A and C.

The domain swapping can be observed from the side following the backbone of each chain as well: A chain, B chain, C chain, and D chain. And all for comparison.

• The symmetry is an overall four-fold for the TMD. Thus, remarkably, the symmetry switches from an overall two-fold symmetry for the ATD and LBD to four-fold for the TMD.

As a result of the swapping and symmetry mismatch, there is subunit non-equivalence; even though all the chains are the same chemically, there are 2 distinct conformations of the subunits. This means there are 2 matching pairs of subunits.

• A is equivalent to C
• B is equivalent to D
• Subunit A is equivalent to Subunit C (in the small structure window in this section). In the main window, a morph showing the equivalency of the 2 subunits by rotating around the axis of their symmetry.
• Subunit B is equivalent to Subunit D (in the small structure window in this section). In the main window, a morph showing the equivalency of the two subunits by rotating around the axis of their symmetry.

However, each of the subunit A/C group though is distinct from those of the B/D group. Having established the two equivalent groups we can simplify the discussion of the relationship between the two pairs by focusing solely on comparing Subunit A' and Subunit B.
The domains themselves stay relatively static between the two conformational forms, with the linkers in between and the resulting arrangement changing. This is best illustrated by superposition of the individual domains of Subunit A and Subunit B:

• Superposition of the ATD.
• Superposition of the LBD.
• Superposition of the TMD.

Subunit A morphing into Subunit B best illustrates how portions, especially the linkers, of the protein change between the two conformational forms.

The linkers are key; besides playing roles in domain swapping and resolving the symmetry mismatch, they are also responsible for relaying the modulation signals from the ATD to the other domains and signaling the conformational change of the LBD to control the opening and closing of the pore. Beyond the two conformations seen here though this particular structure (3kg2) of the receptor does not shed light on the transduction process.

Transmembrane domain architecture and the occluded pore

• Transmembrane segments M1 to M4 depicted in different colors to show the approximate 4-fold rotational symmetry of the entire ion channel domain.

• M1
• M2
• M3
• M4

• The segments shown again, this time parallel to the 4-fold axis.

There is no pore visible in the center consistent with the channel being in a closed state with the antagonist ZK200775 bound to the LBD.

It is the tight helix crossing of specifically the M3 helices that occludes the channel. [BE PATIENT as a small surface is generated.]

Note the differences between the conformations of the carboxy-termini ('top') of the subunit A/C and B/D M3 segments. This is in part is why the symmetry is only approximately four-fold and is one of the several intriguing observations in regard to symmetry for this macromolecule. In fact, the location of 2-fold symmetry at the ends of M3 is just above the portion that spans the membrane and is close to the last region of the structure that doesn't show four-fold symmetry as abruptly below this point everything is 4-fold symmetric.

• To better observe the contributions of each of the membrane segments to the subunit-subunit interactions, the transmembrane domains of three subunits are shown in a surface representation with the segments M1-M4 of the fourth subunit shown as green cylinders. [Note: this scene generates a substantial surface which may take about a minute to calculate. Be patient.]

Note that the M4 segment associates with the ion-channel core of an adjacent subunit.

Fancy, high-quality cartoons on/off.

• The TMD domain of the GluA2 receptor shares structural and sequence similarity with the pore region of the potassium (K+), as hinted at by earlier work^[9][10][11]. Here the pore region of Streptomyces lividans potassium channel (1bl8) superposed with the TMD domain of GluA2, specifically the inner helix of the K+ channel aligned with the M3 segment. The M1 segment of GluA2 also overlays well with the outer helix of the K+ channel even though these portions weren't even included in the calculation of the alignment seen here.

ATP-gated P2X receptor cation channel family

Examples: 3h9v, 3i5d

PIP₂-gated channels

Phosphatidylinositol 4,5-bisphosphate (PIP₂) binds to and directly activates inwardly rectifying potassium channels. Inward rectifier KCh:

See Potassium channel 3D structures