Structural insights into the competitive inhibition of the ATP-gated P2X receptor channel

Kasuya, Go; Yamaura, Toshiaki; Ma, Xiaobo; Nakamura, Ryoki; Takemoto, Mizuki; Nagumo, Hiromitsu; Tanaka, Eiichi; Dohmae, Naoshi; Nakane, Takanori; Yu, Ye; Ishitani, Ryuichiro; Matsuzaki, Osamu; Hattori, Motoyuki; Nureki, Osamu

doi:10.1038/s41467-017-00887-9

Cited by 86 publications

(83 citation statements)

References 53 publications

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“…We therefore inferred that the activation of P2 receptors may be involved in ETX-induced death of MDCK cells. The efflux of K + , and the influx of Na + and Ca 2+ , are likely the result of activation of P2X receptors since P2X receptors are non-selective cation channels [40]. However, antagonists of P2 receptors did not have any effect on the inhibition of ETX-induced death in MDCK cells (S7 Fig).…”

Section: Discussionmentioning

confidence: 99%

Hemolysis in human erythrocytes by Clostridium perfringens epsilon toxin requires activation of P2 receptors

et al. 2018

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Epsilon-toxin (ETX) is produced by types B and D strains of Clostridium perfringens, which cause fatal enterotoxaemia in sheep, goats and cattle. Previous studies showed that only a restricted number of cell lines are sensitive to ETX and ETX-induced hemolysis has not previously been reported. In this study, the hemolytic ability of ETX was examined using erythrocytes from 10 species including murine, rabbit, sheep, monkey and human. We found that ETX caused hemolysis in human erythrocytes (HC50 = 0.2 μM) but not erythrocytes from the other test species. Moreover, the mechanism of ETX-induced hemolysis was further explored. Recent studies showed that some bacterial toxins induce hemolysis through purinergic receptor (P2) activation. Hence, the function of purinergic receptors in ETX-induced hemolysis was tested, and we found that the non-selective P2 receptor antagonists PPADS inhibited ETX-induced lysis of human erythrocytes in a concentration-dependent manner, indicating that ETX-induced hemolysis requires activation of purinergic receptors. P2 receptors comprise seven P2X (P2X1–7) and eight P2Y (P2Y1, P2Y2, P2Y4, P2Y6, and P2Y11–P2Y14) receptor subtypes. The pattern of responsiveness to more selective P2-antagonists implies that both P2Y13 and P2X7 receptors are involved in ETX-induced hemolysis in human species. Furthermore, we demonstrated that extracellular ATP is likely not involved in ETX-induced hemolysis and the activation of P2 receptors. These findings clarified the mechanism of ETX-induced hemolysis and provided new insight into the activities and ETX mode of action.

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Section: Discussionmentioning

confidence: 99%

Hemolysis in human erythrocytes by Clostridium perfringens epsilon toxin requires activation of P2 receptors

et al. 2018

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“…Protonation states for each residue were assigned using PDB2PQR and PROPKA 3.0 (Dolinsky et al, 2004). The modal highest-scoring pose from the docking run was selected (PDB accession #5XW6, (Kasuya et al, 2017)) and distances were measured from a pseudo atom at the centre of the fluorescent moiety. TNP-ATP (PDB #3AR7, (Toyoshima et al, 2011)) was positioned into the first nucleotide binding domain of SUR1 (PDB #6PZI, (Martin et al, 2019)) using the alignment tool in Pymol (Schrödinger, LLC; New York, NY).…”

Section: Surface Expression Assaysmentioning

confidence: 99%

Nucleotide inhibition of the pancreatic ATP-sensitive K+ channel explored with patch-clamp fluorometry

Usher

Ashcroft

Puljung

2019

Preprint

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Pancreatic ATP-sensitive K + channels (KATP) comprise four inward rectifier subunits (Kir6.2), each associated with a sulphonylurea receptor (SUR1). ATP/ADP binding to Kir6.2 shuts KATP.Mg-nucleotide binding to SUR1 stimulates KATP. In the absence of Mg 2+ , SUR1 increases the apparent affinity for nucleotide inhibition at Kir6.2 by an unknown mechanism. We simultaneously measured channel currents and nucleotide binding to Kir6.2. Fits to combined data sets suggest that KATP closes with only one nucleotide molecule bound. A Kir6.2 mutation (C166S) that increases channel activity did not affect nucleotide binding, but greatly perturbed the ability of bound nucleotide to inhibit KATP. Mutations at position K205 in SUR1 affected both nucleotide affinity and the ability of bound nucleotide to inhibit KATP. This suggests a dual role for SUR1 in KATP inhibition, both in directly contributing to nucleotide binding and in stabilising the nucleotidebound closed state.

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“…In the open-channel form of the P2X4 channel of the Golf Coast tick, no binding is observed in the open channel, as the carbonyl group of the residue at the pore constriction, Val361, is hydrogen bonded within helix TM2 (38). For the P2X7 channel bound to the competitive antagonist TNP-ATP, the channel is in an expanded, incompletely activated conformation (39), and cholesterol does not bind in the pore but to ÀOH containing residues exposed on the protein surface (Table S2). The acid-sensing ion channel adopts a structure very similar to those of the P2X channels (40), and the closed form of the acid-sensing ion channel shows no binding of cholesterol either in the pore or to surface exposed sites.…”

Section: Channelsmentioning

confidence: 99%

A Database of Predicted Binding Sites for Cholesterol on Membrane Proteins, Deep in the Membrane

Lee

2018

Biophysical Journal

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The outer membranes of animal cells contain high concentrations of cholesterol, of which a small proportion is located deep within the hydrophobic core of the membrane. An automated docking procedure is described that allows the characterization of binding sites for these deep cholesterol molecules on the membrane-spanning surfaces of membrane proteins and in protein cavities or pores, driven by hydrogen bond formation. A database of this class of predicted binding site is described, covering 397 high-resolution structures. The database includes sites on the transmembrane surfaces of many G-protein coupled receptors; within the fenestrations of two-pore K þ channels and ATP-gated P2X3 channels; in the central cavities of a number of transporters, including Glut1, Glut5, and P-glycoprotein; and in deep clefts in mitochondrial complexes III and IV.

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Structural insights into the competitive inhibition of the ATP-gated P2X receptor channel

Cited by 86 publications

References 53 publications

Hemolysis in human erythrocytes by Clostridium perfringens epsilon toxin requires activation of P2 receptors

Hemolysis in human erythrocytes by Clostridium perfringens epsilon toxin requires activation of P2 receptors

Nucleotide inhibition of the pancreatic ATP-sensitive K+ channel explored with patch-clamp fluorometry

A Database of Predicted Binding Sites for Cholesterol on Membrane Proteins, Deep in the Membrane

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