Atomic distance estimates from disulfides and high-affinity metal-binding sites in a K+ channel pore

Krovetz, Howard S.; VanDongen, Hendrika M.A.; VanDongen, Antonius M.J.

doi:10.1016/s0006-3495(97)78651-x

Cited by 45 publications

(51 citation statements)

References 32 publications

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“…This high-affinity block requires the ␣S583C mutations on the two ␣ENaC subunits of the heteromultimeric channel. If the ␣ENaC subunits face each other across the channel pore as suggested by a study using concatameric constructs (Firsov et al, 1998), this would imply that the optimal distance for coordinated ligation of Zn 2ϩ of ϳ5 Å (Krovetz et al, 1997) would correspond to the distance between the two ␣S583 residues across the pore.…”

Section: Discussionmentioning

confidence: 99%

Mutations in the Epithelial Na⁺ Channel ENaC Outer Pore Disrupt Amiloride Block by Increasing Its Dissociation Rate

Kellenberger

Gautschi²,

Schild³

2003

Mol Pharmacol

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The epithelial Na ϩ channel ENaC mediates transepithelial Na ϩ transport in the distal kidney, the colon, and the lung and is a key element for the maintenance of Na ϩ balance and the regulation of blood pressure. Mutagenesis studies have identified residues ␣S583 and the homologous ␤G525 and ␥G537 in the outer pore entrance that are critical for ENaC block by the K ϩ -sparing diuretic amiloride. The aim of the present study was to determine first, whether these residues are part of the amiloride binding site, and second, whether they are general determinants of ENaC block by amiloride and its derivatives. Kinetic analysis of the association and dissociation rates of amiloride and benzamil to ENaC showed that mutation of residue ␣S583C and the homologous ␤G525C increased the dissociation rate of the drugs from the binding site, with little changes in their association rate. Thus, these mutations destabilize the binding interaction between the blockers and the receptor on the channel, favoring the unbinding of the ligand. This strongly suggests that they are part of the binding site. Because mutations of ␣S583, ␤G525, and ␥G537 have similar effects on amiloride, benzamil, and triamterene block, we conclude that these three ENaC blockers share a common receptor within the ion channel pore.

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Section: Discussionmentioning

confidence: 99%

Mutations in the Epithelial Na⁺ Channel ENaC Outer Pore Disrupt Amiloride Block by Increasing Its Dissociation Rate

Kellenberger

Gautschi²,

Schild³

2003

Mol Pharmacol

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“…Crystallographic and mutagenesis studies on metalloproteins and channels reveal some of the details of the coordination of metal cations by sulfhydryls (26,28,31,32). Cd 2ϩ is a soft metal cation (Lewis acid) that preferentially interacts with soft bases such as deprotonated sulfhydryl groups (S Ϫ ) (29).…”

Section: Discussionmentioning

confidence: 99%

Minimal Structural Rearrangement of the Cytoplasmic Pore during Activation of the 5-HT3A Receptor

Panicker

Cruz

Arrabit

et al. 2004

Journal of Biological Chemistry

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Ligand-gated ion channel receptors mediate the response of fast neurotransmitters by opening in less than a millisecond. Here, we investigated the activation mechanism of a serotonin-gated receptor (5-HT 3A ) by systematically introducing cysteine substitutions throughout the pore-lining M1-M2 loop and M2 transmembrane domain. We hypothesized that multiple cysteines in the narrowest region of the pore, which together can form a high affinity binding site for metal cations, would reveal changes in pore structure during gating. Using cadmium (Cd 2؉ ) as a probe, two cysteine substitutions in the cytoplasmic selectivity filter, S2 C and, to a lesser extent, G-2 C, showed high affinity inhibition with Cd 2؉ Ligand-gated ion channel (LGIC) 1 receptors are responsible for rapid chemical transmission between neurons in the nervous system (1). Alterations in the response of ligand-gated receptors have profound effects on neuronal activity in the brain. For example, mutations in nicotinic acetylcholine-gated receptors (nAChRs) and glycine-gated receptors have been linked to certain forms of epilepsy and startle disease, respectively (2-4).LGIC receptors that respond to acetylcholine, ␥-aminobutyric acid, glycine, or serotonin (5-HT) possess a conserved pair of extracellular cysteines in the N-terminal domain ("Cys loop" receptors) and are composed of five subunits, each containing four putative transmembrane domains (5). Of these four domains, the second transmembrane domain (M2) lines the majority of the water-filled pore (Fig. 1A).The rapid response of LGIC receptors is achieved by the energetic coupling of agonist binding in the extracellular Nterminal domain with a "gate" located 60 Å away in the receptor pore (M2 domain) (6). Several studies with LGIC receptors suggest that the gate is situated in the middle of the M2 (6 -12) (but see Ref. 13). The primary determinants of selectivity appear to be located below the gate, near the cytoplasmic membrane surface. Mutations in the cytoplasmic end of the M2 alter ion selectivity (14 -17), and several recent studies show that by mutating sites in this region, cation-conducting LGIC can be converted to anion-conducting channels and vice versa (18 -23). Because permeability of like-charged molecules in LGIC appears to be largely dependent on size, it is believed that the narrowest region of the open LGIC pore coincides with the channel selectivity filter, which is formed by a portion of the M1-M2 loop and the cytoplasmic side of the M2 helix (16,24). Consistent with this region being narrow, site-specific mutations in the filter alter conductivity in a manner that depends upon side-chain volume (8,24). Thus, the pore of the open receptor can be envisaged as a funnel, with a wide extracellular vestibule that tapers to a constriction at the cytoplasmic side of the membrane, where ion selectivity is determined (25).What is known about the extent of movement in the cytoplasmic selectivity filter? The 9-Å three-dimensional structures of open and closed Torpedo nAChRs derived fro...

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“…In contrast to SCAM, disulfide cross-linking experiments test the ability of two engineered Cys residues to form a disulfide bond (25)(26)(27). The rate of disulfide bond formation depends on collision frequency between the pair of Cys residues, which in turn depends on the proximity and relative mobility of the two protein regions.…”

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confidence: 99%

Gating-induced Conformational Rearrangement of the γ-Aminobutyric Acid Type A Receptor β-α Subunit Interface in the Membrane-spanning Domain

Bali

Akabas

2012

Journal of Biological Chemistry

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Background: GABA A receptor transmembrane domain undergoes conformational changes during channel gating. Results: Solvent accessibility increases in a cluster of ␣1 subunit M1 segment residues facing the ␤2 subunit M3 segment during channel gating. Conclusion: Channel gating induces a conformational change in the subunit interface between ␣1M1 and ␤2M3 and within the ␣1 subunit. Significance: This work increases understanding of pentameric ligand-gated ion channel gating.

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Atomic distance estimates from disulfides and high-affinity metal-binding sites in a K+ channel pore

Cited by 45 publications

References 32 publications

Mutations in the Epithelial Na⁺ Channel ENaC Outer Pore Disrupt Amiloride Block by Increasing Its Dissociation Rate

Mutations in the Epithelial Na⁺ Channel ENaC Outer Pore Disrupt Amiloride Block by Increasing Its Dissociation Rate

Minimal Structural Rearrangement of the Cytoplasmic Pore during Activation of the 5-HT3A Receptor

Gating-induced Conformational Rearrangement of the γ-Aminobutyric Acid Type A Receptor β-α Subunit Interface in the Membrane-spanning Domain

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Atomic distance estimates from disulfides and high-affinity metal-binding sites in a K+ channel pore

Cited by 45 publications

References 32 publications

Mutations in the Epithelial Na+ Channel ENaC Outer Pore Disrupt Amiloride Block by Increasing Its Dissociation Rate

Mutations in the Epithelial Na+ Channel ENaC Outer Pore Disrupt Amiloride Block by Increasing Its Dissociation Rate

Minimal Structural Rearrangement of the Cytoplasmic Pore during Activation of the 5-HT3A Receptor

Gating-induced Conformational Rearrangement of the γ-Aminobutyric Acid Type A Receptor β-α Subunit Interface in the Membrane-spanning Domain

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Mutations in the Epithelial Na⁺ Channel ENaC Outer Pore Disrupt Amiloride Block by Increasing Its Dissociation Rate

Mutations in the Epithelial Na⁺ Channel ENaC Outer Pore Disrupt Amiloride Block by Increasing Its Dissociation Rate