The structural patterns of the potentiation and the blockade of inhibitory cys-loop receptors through the transmembrane domain

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Abstract

Anion-conducting cys-loop receptors activated by γ-aminobutyric acid (GABAАRs) and glycine (GlyRs) have inhibitory activity in the brain and spinal cord. GABAАRs and GlyRs are targets for various substances that potentiate or inhibit the receptor functions. Many of these substances are clinically significant agents to treat neurological and psychiatric conditions.

The review covers both our results and literature data on electrophysiology, mutations, and biochemistry of non-competitive antagonists, general anesthetics, barbiturates, and fenamates modulating GABAАRs and GlyRs. We focused on our own molecular modeling to determine the sites and the characteristics of binding of these substances to the GABAАR and GlyR transmembrane domain. With the structural patterns of the binding, we have identified possible molecular mechanisms of action for these substances.

About the authors

Alexey V. Rossokhin

Research Center of Neurolgy

Author for correspondence.
Email: alrossokhin@yandex.ru
ORCID iD: 0000-0001-7024-7461

Cand. Sci. (Phys.-Math.), leading researcher, Laboratory of functional synaptology, Brain Institute

Russian Federation, 105064, Moscow, Obukha per., 5

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Supplementary files

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2. Fig. 1. Ligand-activated pentamer receptor architecture. А — the view of the receptor in the membrane plane. The frontal subunit is omitted to demonstrate a pore. The arrow indicates the receptor central axis. The transmembrane domain (TMD) and the extracellular domain (ECD) are pointed out; B — the extracellular view of TMD. Transmembrane chains М1–М4 are indicated on the colored subunit. Intersubunit interfaces are pointed with + and –; C — the alignment of GABAАR and GlyR segment M1–M3 aminoacid sequences. The GABAАR_α1 P14867, GABAАR_β1 P18505, GABAАR_β2 P47870, GABAАR_γ2 P18507, GlyR_α1 P23415, GlyR_α2 P23416, and GlyR_β P48167 sequences were sourced from the UniProt Database and aligned relative to the highly conserved Arg 0' residues. Intersubunit potentiating site residues are marked with numbers and strokes above the sentences.

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3. Fig. 2. GABAАR pore. А — visualized space inside the pore of the α1β2γ2 GABAАR models that are homologous to the closed α3 GlyR (5CFB), open α1 GlyR (3JAE), and desensitized β3 GABAАR (4COF) structures. Only М2 segments are depicted. The frontal subunit was omitted for clarity. Horizontal lines indicate residue levels -2′, 9′, and 20′ for main pore constrictions. Some subunits are highlighted in cyan (α1), orange (β2), and purple (γ2); В — dependence of the pore diameter on the pore depth in the closed, open, and desensitized receptor states. Vertical lines indicate pore levels -2′, 9′, and 20′.

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4. Fig. 3. GABAАR non-competitive antagonists. А — cryo-EM structure of GABAАR (6X40) complexed with picrotoxin; B, C — binding of penicillin and furosemide in the pore of the GABAАR model that is homologous to the bacterial receptor GLIC (3HEZ) structure. A–C present views from the extracellular space (top) and in the plane of membrane (bottom). The frontal subunit is omitted for clarity on the side-view images. The images show side chains of residues 2′, 6′, 9′, and 13′ that most significantly contribute to the blocker-receptor interaction. Hydrogen bonds are depicted as red dashed lines. Color subunit presentation complies with Fig. 2.

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5. Fig. 4. Fenamate GABAАR potentiation and inhibition. The potentiating sites of binding meclofenamic acid (MFA; А) and niflumic acid (NFA; В) in β(+)/α(−) transmembrane interface of the α1β2γ2 GABAАR model that is homologous to the 3JAE α1 GlyR structure. C — superposition of the α1β1γ2 и α1β2γ2 GABAАR models in the β(+)/α(−) interface region. R19′ side chain in β2 subunit is green; D — NFA energy profile in the pore of the α1β2γ2 GABAАR model; E — structural models of NFA binding in the pore at the upper and lower sites. А, B, and E present residue side chains that significantly contribute (>1 kcal/mol) to the energy of ligand-receptor interaction. Hydrogen bonds are depicted as red dashed lines. Color presentation of subunits complies with Fig. 2.

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6. Fig. 5. Structural determinants of GlyR potentiation. А, B — TMD of α1 GlyR in open (3JAE) and closed (3JAD) states. The images present residue side chains R19′ and Q(–26′). 3JAE and 3JAD structures are shown as they are laid out in Protein Data Bank without preliminary energy minimization. С–Е — view of the transmembrane β(+)/α(−) (С), α(+)/β(−) (D), and β(+)/β(−) (E) interfaces of the α1β GlyR model in the membrane plane that are homologous to the 3JAE structure. The figure presents the parts of the M1–M3 helices and the residue side chains that are homologous to those involved in MFA binding on the β(+)/α(−) interface of α1β2γ2 GABAАR. Hydrogen bonds are depicted as red dashed lines. Some subunits are highlighted in cyan (α1) and orange (β).

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