Structural Basis for High-Affinity Trapping of the NaV1.7 Channel in Its Resting State by Tarantula Toxin

Wisedchaisri, Goragot; Tonggu, Lige; El-Din, Tamer M. Gamal; McCord, E.; Zheng, Ning; Catterall, William A.

doi:10.1016/j.molcel.2020.10.039

Cited by 54 publications

(87 citation statements)

References 72 publications

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“…We performed docking experiments with Tap1a-WT and Tap1a-OPT1 on these available channel structures, and the models that produced closer interactions between VSDII and Tap1a used the structure determined for the complex hNa V 1.7 VSDII-ProTx-II ( Xu et al, 2019 ), and the hNa V 1.4 channel structure determined in complex with the β1 auxiliary subunit ( Pan et al, 2018 ). Our findings with these channel structures are consistent with the structure-function relationship findings for other NaSpTx peptides ( Cardoso and Lewis, 2019 ; Shen et al, 2019 ; Xu et al, 2019 ; Mueller et al, 2020 ; Wisedchaisri et al, 2021 ).…”

Section: Discussionsupporting

confidence: 90%

“…In Tap1a-OPT1, h-bonds were formed by K31 and the new K33 in the C-terminal. Similarly, a study with the optimized peptide m3-HwTx-IV and the Na V 1.7 performed via Cryo-EM revealed K27 in loop 4 and K32 at the C-terminal as the main drivers of this inhibitory mechanism ( Wisedchaisri et al, 2021 ). Besides forming strong h-bonds, lysin residues can effectively interact with lipids in the cell membrane as demonstrated in studies with the peptide ProTx-II ( Henriques et al, 2016 ).…”

Section: Discussionmentioning

confidence: 97%

“…Although Cryo-EM and X-ray studies of Na V channels have disclosed their three-dimensional structure in detail, these were mostly performed with channels at the open-state in which the voltage-sensor S4 is upwards and the channel pore open ( Pan et al, 2018 ; Pan et al, 2019 ; Shen et al, 2019 ; Pan et al, 2021 ; Wisedchaisri et al, 2021 ), or did not describe key residues in the loop S3–S4 of VSDII which are essential for peptide binding ( Shen et al, 2019 ). We performed docking experiments with Tap1a-WT and Tap1a-OPT1 on these available channel structures, and the models that produced closer interactions between VSDII and Tap1a used the structure determined for the complex hNa V 1.7 VSDII-ProTx-II ( Xu et al, 2019 ), and the hNa V 1.4 channel structure determined in complex with the β1 auxiliary subunit ( Pan et al, 2018 ).…”

Section: Discussionmentioning

confidence: 99%

See 2 more Smart Citations

Engineering of a Spider Peptide via Conserved Structure-Function Traits Optimizes Sodium Channel Inhibition In Vitro and Anti-Nociception In Vivo

Mawlawi

Zhao

et al. 2021

Front. Mol. Biosci.

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Venom peptides are potent and selective modulators of voltage-gated ion channels that regulate neuronal function both in health and in disease. We previously identified the spider venom peptide Tap1a from the Venezuelan tarantula Theraphosa apophysis that targeted multiple voltage-gated sodium and calcium channels in visceral pain pathways and inhibited visceral mechano-sensing neurons contributing to irritable bowel syndrome. In this work, alanine scanning and domain activity analysis revealed Tap1a inhibited sodium channels by binding with nanomolar affinity to the voltage-sensor domain II utilising conserved structure-function features characteristic of spider peptides belonging to family NaSpTx1. In order to speed up the development of optimized NaV-targeting peptides with greater inhibitory potency and enhanced in vivo activity, we tested the hypothesis that incorporating residues identified from other optimized NaSpTx1 peptides into Tap1a could also optimize its potency for NaVs. Applying this approach, we designed the peptides Tap1a-OPT1 and Tap1a-OPT2 exhibiting significant increased potency for NaV1.1, NaV1.2, NaV1.3, NaV1.6 and NaV1.7 involved in several neurological disorders including acute and chronic pain, motor neuron disease and epilepsy. Tap1a-OPT1 showed increased potency for the off-target NaV1.4, while this off-target activity was absent in Tap1a-OPT2. This enhanced potency arose through a slowed off-rate mechanism. Optimized inhibition of NaV channels observed in vitro translated in vivo, with reversal of nocifensive behaviours in a murine model of NaV-mediated pain also enhanced by Tap1a-OPT. Molecular docking studies suggested that improved interactions within loops 3 and 4, and C-terminal of Tap1a-OPT and the NaV channel voltage-sensor domain II were the main drivers of potency optimization. Overall, the rationally designed peptide Tap1a-OPT displayed new and refined structure-function features which are likely the major contributors to its enhanced bioactive properties observed in vivo. This work contributes to the rapid engineering and optimization of potent spider peptides multi-targeting NaV channels, and the research into novel drugs to treat neurological diseases.

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Section: Discussionsupporting

confidence: 90%

Section: Discussionmentioning

confidence: 97%

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Engineering of a Spider Peptide via Conserved Structure-Function Traits Optimizes Sodium Channel Inhibition In Vitro and Anti-Nociception In Vivo

Mawlawi

Zhao

et al. 2021

Front. Mol. Biosci.

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“…Although the X-ray crystal structure of Kv1.3 is currently not known, a cryo-EM structure of the channel is under review (http:// dx.doi.org/10.2139/ssrn.34672 50). Analysis of the cryo-EM structure of the bacterial NavAB/human Nav1.7 Na + channel chimera in complex with the tarantula toxin Huwentoxin-IV allowed the identification of the key toxin residue that interacts with the voltage-sensor of the channel and stabilize the complex in the resting state (Wisedchaisri et al 2021). This and other cryo-EM structures of the toxin-channel complexes may be extremely useful in guiding AI-based drug design (Lau et al 2018).…”

Section: Artificial Intelligence (Ai)-guided Drug Design and Phage DImentioning

confidence: 99%

The Kv1.3 K+ channel in the immune system and its “precision pharmacology” using peptide toxins

2021

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Since the discovery of the Kv1.3 voltage-gated K+ channel in human T cells in 1984, ion channels are considered crucial elements of the signal transduction machinery in the immune system. Our knowledge about Kv1.3 and its inhibitors is outstanding, motivated by their potential application in autoimmune diseases mediated by Kv1.3 overexpressing effector memory T cells (e.g., Multiple Sclerosis). High affinity Kv1.3 inhibitors are either small organic molecules (e.g., Pap-1) or peptides isolated from venomous animals. To date, the highest affinity Kv1.3 inhibitors with the best Kv1.3 selectivity are the engineered analogues of the sea anemone peptide ShK (e.g., ShK-186), the engineered scorpion toxin HsTx1[R14A] and the natural scorpion toxin Vm24. These peptides inhibit Kv1.3 in picomolar concentrations and are several thousand-fold selective for Kv1.3 over other biologically critical ion channels. Despite the significant progress in the field of Kv1.3 molecular pharmacology several progressive questions remain to be elucidated and discussed here. These include the conjugation of the peptides to carriers to increase the residency time of the peptides in the circulation (e.g., PEGylation and engineering the peptides into antibodies), use of rational drug design to create novel peptide inhibitors and understanding the potential off-target effects of Kv1.3 inhibition.

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“…Cryo-EM structures of several eukaryotic channels in the apo form and in complex with different toxins are now available, e.g., [20][21][22][23][24]. Although these structures are useful in rationalizing numerous mutational and ligand-binding studies, including action of chemical insecticides like pyrethroids and proteinaceous toxins, they are of limited resolution and represent snapshots of certain conformations captured in channel states with presumably inactivated pore domain [25].…”

Section: Introductionmentioning

confidence: 99%

Mapping the interaction surface of scorpion β-toxins with an insect sodium channel

Zhorov

Song

et al. 2021

Biochemical Journal

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The interaction of insect-selective scorpion depressant β-toxins (LqhIT2 and Lqh-dprIT3 from Leiurus quinquestriatus hebraeus) with the Blattella germanica sodium channel, BgNav1-1a, was investigated using site-directed mutagenesis, electrophysiological analyses, and structural modeling. Focusing on the pharmacologically-defined binding site-4 of scorpion β-toxins at the voltage-sensing domain II (VSD-II), we found that charge neutralization of D802 in VSD-II greatly enhanced the channel sensitivity to Lqh-dprIT3. This was consistent with the high sensitivity of the splice variant BgNav2-1, bearing G802, to Lqh-dprIT3, and low sensitivity of BgNav2-1 mutant, G802D, to the toxin. Further mutational and electrophysiological analyses revealed that the sensitivity of the WT = D802E < D802G < D802A < D802K channel mutants to Lqh-dprIT3 correlated with the depolarizing shifts of activation in toxin-free channels. However, the sensitivity of single mutants involving IIS4 basic residues (K4E = WT << R1E < R2E < R3E) or double mutants (D802K = K4E/D802K = R3E/D802K > R2E/D802K > R1E/D802K > WT) did not correlate with the activation shifts. Using the cryo-EM structure of the Periplaneta americana channel, NavPaS, as template and the crystal structure of LqhIT2, we constructed structural models of LqhIT2 and Lqh-dprIT3-c in complex with BgNav1-1a. These models along with the mutational analysis suggest that depressant toxins approach the salt-bridge between R1 and D802 at VSD-II to form contacts with linkers IIS1-S2, IIS3-S4, IIIP5-P1 and IIIP2-S6. Elimination of this salt-bridge enables deeper penetration of the toxin into a VSD-II gorge to form new contacts with the channel, leading to increased channel sensitivity to Lqh-dprIT3.

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Structural Basis for High-Affinity Trapping of the NaV1.7 Channel in Its Resting State by Tarantula Toxin

Cited by 54 publications

References 72 publications

Engineering of a Spider Peptide via Conserved Structure-Function Traits Optimizes Sodium Channel Inhibition In Vitro and Anti-Nociception In Vivo

Engineering of a Spider Peptide via Conserved Structure-Function Traits Optimizes Sodium Channel Inhibition In Vitro and Anti-Nociception In Vivo

The Kv1.3 K+ channel in the immune system and its “precision pharmacology” using peptide toxins

Mapping the interaction surface of scorpion β-toxins with an insect sodium channel

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