Structures of human Na
            <sub>v</sub>
            1.7 channel in complex with auxiliary subunits and animal toxins

Shen, Huaizong; Liu, Dongliang; Wu, Kun; Lei, Jianlin; Yan, Nieng

doi:10.1126/science.aaw2493

Cited by 383 publications

(566 citation statements)

References 67 publications

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“…Icilin/Ion channel TRPM8 (PDB:6NR3, EMD:0487) 41 ; LY2119620/M2R (PDB:6OIK, EMD:20079) 38 ; Iperoxo/M2R (PDB:6OIK, EMD:20079) 38 ; Saxitoxin/Nav1.7 (PDB: 6J8G, EMD:9781) 39 ; Biculine/GABAA (PDB:6HUK, EMD:0280) 37 ; Xanax/GABAA (PDB:6HUO, EMD:0282) 37 ; Valium, GABAA (PDB:6HUP, EMD:0283) 37 ; NDI-091143/ATP Citrate Lyase (PDB:6O0H, EMD:0567) 7 ; paroxetine/serotonin transporter (PDB:6DZW, EMD:8941) 40 ;…”

Section: Methodsmentioning

confidence: 99%

“…Using GlideEM we were able to identify candidate poses in this resolution range with cross correlations that were comparable to the deposited poses for all of the structures that we studied. For GABAA in complex with three different ligands 37 , the M2 muscarinic acetylcholine receptor in complex with two different ligands 38 , the sodium channel Nav1.7 with one ligand 39 , ATP citrate lyase with one ligand 7 , and the serotonin transporter with two different ligands 40 the poses we identified with GlideEM largely agreed with what was deposited in the PDB ( Supplementary Fig. 7) (although it should be noted that for the deposited structures of the M2 receptor and serotonin transporter the ligands were docked with the traditional version of Glide).…”

Section: Using Gemspot With 30-45 å Resolution Mapsmentioning

confidence: 99%

See 1 more Smart Citation

GemSpot: A Pipeline for Robust Modeling of Ligands into CryoEM Maps

Robertson¹,

Zundert

Borrelli

et al. 2019

Preprint

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AbstractProducing an accurate atomic model of biomolecule-ligand interactions from maps generated by cryo-electron microscopy often presents challenges inherent to the methodology and the dynamic nature of ligand binding. Here we have developed GemSpot, a pipeline of computational chemistry methods that take into account EM map potentials, quantum mechanics energy calculations, and water molecule site prediction to generate candidate poses and provide a measure of the degree of confidence. The pipeline is validated through several published cryoEM structures of complexes in different resolution ranges and various types of ligands. In all cases, at least one identified pose produced both excellent interactions with the target and agreement with the map. GemSpot will be valuable for the robust identification of ligand poses and drug discovery efforts through cryoEM.

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Section: Methodsmentioning

confidence: 99%

Section: Using Gemspot With 30-45 å Resolution Mapsmentioning

confidence: 99%

GemSpot: A Pipeline for Robust Modeling of Ligands into CryoEM Maps

Robertson¹,

Zundert

Borrelli

et al. 2019

Preprint

View full text Add to dashboard Cite

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“…Notably, the upper region of the selectivity filter near the KIIIA binding site has a minimum radius of ~2.5 Å, which is large enough to potentially allow sodium ion conduction and consistent with the characteristic incomplete block of Nav channels by KIIIA (13,15). Notably, KIIIA is positioned just above the selectivity filter which is different from small molecular toxins, such as TTX and STX, which are positioned deeper into the selectivity region (25,26).…”

Section: Molecular Modeling Reveals Asymmetric Binding Of Kiiia To Hnmentioning

confidence: 83%

“…Another feature of the molecular mechanism of KIIIA binding are differences in the reversibility and irreversibility of some of the KIIIA mutants on different Nav channels. Cryo-EM structures of hNav1.2, hNav1.4, and hNav1.7 channels (25,31,32) revealed that the extracellular vestibule of the channel pore targeted by KIIIA is surrounded by several relatively long loop regions raising the possibility that KIIIA access pathway to the toxin binding site is relatively narrow. Restricted access and escape pathways for KIIIA binding agree with the relatively slow kon and koff rates observed in previously published data (2,13,15) and our functional studies (Figures 2 and 3).…”

Section: Discussionmentioning

confidence: 99%

Molecular Determinants of μ-Conotoxin KIIIA Interaction with the Human Voltage-Gated Sodium Channel NaV1.7

Kimball¹,

Nguyen²,

Olivera

et al. 2019

Preprint

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The voltage-gated sodium (Nav) channel subtype Nav1.7 plays a critical role in pain signaling, making it an important drug target. A number of peptide toxins from cone snails (conotoxins) bind to the extracellular vestibule of the Nav channel pore and block ion conduction. While the known conotoxins have variable selectivity among Nav channel subtypes, they form potential scaffolds for engineering of selective and potent channel inhibitors. Here we studied the molecular interactions between μ-conotoxin KIIIA (KIIIA) and the human Nav1.7 channel (hNav1.7). Using the cryo-electron microscopy (cryo-EM) structure of the electric eel Nav1.4 channel as a template we developed a structural model of hNav1.7 with Rosetta computational modeling. We performed in silico docking of KIIIA using RosettaDock and identified residues forming specific pairwise contacts between KIIIA and hNav1.7. Pairwise interactions were experimentally validated using mutant cycle analysis. Comparison with a recently published cryo-EM structure of the KIIIA-hNav1.2 channel complex revealed key similarities and differences between channel subtypes with potential implications for the molecular mechanism of toxin block. Our integrative approach, combining high-resolution structural data with computational modeling and experimental validation, will be useful for engineering of molecular probes to study Nav channels function and for rational design of novel biologics to treat chronic pain, cardiac arrhythmias, and epilepsy.

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“…Structural models of human SCN5A (UniProtKB accession number: Q14524-1, modeled residues: 30-440, 685-957, 1174-1887) bound with SCN1B (UniProtKB accession number: Q07699-1, modeled residues: 20-192) were generated by homology modeling using the protein structure prediction software Rosetta (version 3.10). 31 The cryo-EM structure of human SCN9A bound with SCN1B and the Ig domain of SCN2B resolved to 3.2 Å (PDB 6J8H) 32 was used as the primary template while the cryo-EM structure of NavPaS from American Cockroach resolved to 2.6 Å (PDB ID: 6A95) 33 Methods and Table S4.…”

Section: Homology Model and Structural Calculationsmentioning

confidence: 99%

High-throughput reclassification ofSCN5Avariants

Glazer

Wada

Bian

et al. 2019

Preprint

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Rationale: Partial or complete loss of function variants in SCN5A are the most common genetic cause of the arrhythmia disorder Brugada Syndrome (BrS1). However, the pathogenicity of SCN5A variants is often unknown or disputed; 80% of the 1,390 SCN5A missense variants observed in at least one individual to date are variants of uncertain significance (VUS). The designation of VUS is a barrier to the use of sequence data in clinical care. Objective:We selected 83 variants for study: 10 previously studied control variants, 10 suspected benign variants, and 63 suspected Brugada Syndrome-associated variants, selected on the basis of their frequency in the general population and in patients with Brugada Syndrome. We used high-throughput automated patch clamping to study the function of the 83 variants, with the goal of reclassifying variants with functional data.Methods and Results: Ten previously studied variants had functional properties concordant with published manual patch clamp data. All 10 suspected benign variants had wildtype-like function. 22 suspected BrS variants had loss of channel function (<10% normalized peak current) and 23 variants had partial loss of function (10-50% normalized peak current). The 73 previously unstudied variants were initially classified as likely benign (n=2), likely pathogenic (n=11), or VUS (n=60). After the patch clamp studies, 16 variants were benign/likely benign, 47 were pathogenic/likely pathogenic, and only 10 were still VUS. 8/22 loss of function variants were partially rescuable by incubation at lower temperature or pretreatment with a sodium channel blocker. Structural modeling identified likely mechanisms for loss of function including altered thermostability, and disruptions to alpha helices, disulfide bonds, or the permeation pore. Conclusions:High-throughput automated patch clamping enabled the reclassification of the majority of tested VUS's in SCN5A.

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Structures of human Na _v 1.7 channel in complex with auxiliary subunits and animal toxins

Cited by 383 publications

References 67 publications

GemSpot: A Pipeline for Robust Modeling of Ligands into CryoEM Maps

GemSpot: A Pipeline for Robust Modeling of Ligands into CryoEM Maps

Molecular Determinants of μ-Conotoxin KIIIA Interaction with the Human Voltage-Gated Sodium Channel NaV1.7

High-throughput reclassification ofSCN5Avariants

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Structures of human Na v 1.7 channel in complex with auxiliary subunits and animal toxins

Cited by 383 publications

References 67 publications

GemSpot: A Pipeline for Robust Modeling of Ligands into CryoEM Maps

GemSpot: A Pipeline for Robust Modeling of Ligands into CryoEM Maps

Molecular Determinants of μ-Conotoxin KIIIA Interaction with the Human Voltage-Gated Sodium Channel NaV1.7

High-throughput reclassification ofSCN5Avariants

Contact Info

Product

Resources

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Structures of human Na _v 1.7 channel in complex with auxiliary subunits and animal toxins