Studies Examining the Relationship between the Chemical Structure of Protoxin II and Its Activity on Voltage Gated Sodium Channels

Park, Jae H.; Carlin, Kevin P.; Wu, Gang; Ilyin, Victor I.; Musza, László; Blake, Paul R.; Kyle, Donald J.

doi:10.1021/jm500687u

Cited by 46 publications

(63 citation statements)

References 37 publications

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“…FLIPR studies (Fig. 7A) revealed that synthetic ProTx-II inhibits hNa V 1.7 in SH-SY5Y cells with an IC 50 of 0.40 Ϯ 0.07 nM, which is within the previously determined range (16,29,40). All ProTx-II analogues with mutations in Trp residues had reduced potency against hNav1.7 compared with ProTx-II.…”

Section: Lipidsupporting

confidence: 68%

“…We were specifically interested in examining whether the hydrophobic patch might be important for membrane binding. Previous mutagenesis studies have shown that Trp residues affect the potency (39) and/or the selectivity (40) of ProTx-II as shown with analogues with the single mutations as follows: W5A, W7A, W24L, W30A (39), W30I, or amidated Trp-30 (40). In these earlier studies, the hydrophobic aromatic Trp residues were replaced with nonaromatic amino acid residues that possess a lower tendency to bind/insert into the membrane (41).…”

Section: Protx-ii Interaction With Model Lipidmentioning

confidence: 99%

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Interaction of Tarantula Venom Peptide ProTx-II with Lipid Membranes Is a Prerequisite for Its Inhibition of Human Voltage-gated Sodium Channel NaV1.7

Henriques

Deplazes

Lawrence

et al. 2016

Journal of Biological Chemistry

135

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ProTx-II is a disulfide-rich peptide toxin from tarantula venom able to inhibit the human voltage-gated sodium channel 1.7 (hNa V 1.7), a channel reported to be involved in nociception, and thus it might have potential as a pain therapeutic. ProTx-II acts by binding to the membrane-embedded voltage sensor domain of hNa V 1.7, but the precise peptide channel-binding site and the importance of membrane binding on the inhibitory activity of ProTx-II remain unknown. In this study, we examined the structure and membrane-binding properties of ProTx-II and several analogues using NMR spectroscopy, surface plasmon resonance, fluorescence spectroscopy, and molecular dynamics simulations. Our results show a direct correlation between ProTx-II membrane binding affinity and its potency as an hNa V 1.7 channel inhibitor. The data support a model whereby a hydrophobic patch on the ProTx-II surface anchors the molecule at the cell surface in a position that optimizes interaction of the peptide with the binding site on the voltage sensor domain. This is the first study to demonstrate that binding of ProTx-II to the lipid membrane is directly linked to its potency as an hNa V 1.7 channel inhibitor.Voltage-gated ion channels (VGICs) 4 are transmembrane proteins responsible for voltage-dependent movement of ions across cell membranes. They are involved in a wide range of physiological processes, including action potential generation in excitable cells, muscle and nerve relaxation, regulation of blood pressure, and sensory transduction. Many disorders are associated with VGIC abnormalities, and hence VGICs are actively pursued as drug targets for the treatment of a range of neuromuscular, neurological, or inflammatory disorders (1-3). For instance, the human voltage-gated sodium channel subtype 1.7 (hNa V 1.7) is involved in pain sensation, and molecules that selectively inhibit this channel might therefore be useful as leads for the development of novel analgesics (4). However, high sequence identity and structural homology exist between the nine subtypes of voltage-gated sodium channels, and given the critical role of Na V channels in the normal electrical activity of neurons, skeletal muscles, and cardiomyocytes, subtype selectivity is crucial but often difficult to achieve with small molecule inhibitors (2).Disulfide-rich peptide toxins isolated from animal venoms (e.g. spiders, snakes, and cone snails) have attracted much attention as potential analgesics because of their well defined three-dimensional structure and ability to inhibit voltage-gated sodium (Na V ), potassium (K V ), and calcium (Ca V ) ion channels with high potency and selectivity (5-9). Because of their stability, selectivity, and potency, these disulfide-rich peptides have been extensively characterized and are vigorously being pursued as drug leads as well as pharmacological tools to characterize VGICs (5).In general, peptide toxins that inhibit VGICs can be divided into two main groups based on their inhibitory strategy as fol-

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Section: Lipidsupporting

confidence: 68%

Section: Protx-ii Interaction With Model Lipidmentioning

confidence: 99%

Interaction of Tarantula Venom Peptide ProTx-II with Lipid Membranes Is a Prerequisite for Its Inhibition of Human Voltage-gated Sodium Channel NaV1.7

Henriques

Deplazes

Lawrence

et al. 2016

Journal of Biological Chemistry

135

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“…Loss of function mutations on SCN9A, the gene encoding hNa V 1.7, result in unresponsiveness to pain, whereas gain of function mutations result in increased chronic and acute pain sensations [3,4]. Because of this link to pain, efforts to identify potent and selective inhibitors of hNa V 1.7 are underway [5][6][7][8].…”

Section: Introductionmentioning

confidence: 99%

Spider peptide toxin HwTx-IV engineered to bind to lipid membranes has an increased inhibitory potency at human voltage-gated sodium channel hNa V 1.7

Agwa¹,

Lawrence²,

Deplazes³

et al. 2017

Biochimica et Biophysica Acta (BBA) - Biomembranes

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“…Several studies were undertaken recently with the aim of improving potency, selectivity, stability, and bioavailability of venom-derived lead peptide compounds targeting Nav1.7 (19,(21)(22)(23).…”

Section: The Atomic Coordinates and Structure Factors (Code 5epm) Havmentioning

confidence: 99%

“…CcoTx1, like other site 4 microproteins (binding to S3-S4 linker of domain II) (36 -38), can be effectively dislodged from its binding site on the Nav1.7 channel by strong depolarization (23,44). In our studies, 5 nM D1Ia induced almost complete block of Nav1.7 current after a 20-min application; however, a subsequent positive depolarizing pulse to ϩ50 mV for 1 min has reversed the blocking effect of D1Ia by 83 Ϯ 10% (n ϭ 5) (supplemental Fig.…”

Section: Effects Of Post-translational Modification On Potency Andmentioning

confidence: 99%

Engineering Highly Potent and Selective Microproteins against Nav1.7 Sodium Channel for Treatment of Pain

Shcherbatko

Rossi

Foletti

et al. 2016

Journal of Biological Chemistry

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The prominent role of voltage-gated sodium channel 1.7 (Nav1.7) in nociception was revealed by remarkable human clinical and genetic evidence. Development of potent and subtypeselective inhibitors of this ion channel is crucial for obtaining therapeutically useful analgesic compounds. Microproteins isolated from animal venoms have been identified as promising therapeutic leads for ion channels, because they naturally evolved to be potent ion channel blockers. Here, we report the engineering of highly potent and selective inhibitors of the Nav1.7 channel based on tarantula ceratotoxin-1 (CcoTx1). We utilized a combination of directed evolution, saturation mutagenesis, chemical modification, and rational drug design to obtain higher potency and selectivity to the Nav1.7 channel. The resulting microproteins are highly potent (IC 50 to Nav1.7 of 2.5 nM) and selective. We achieved 80-and 20-fold selectivity over the closely related Nav1.2 and Nav1.6 channels, respectively, and the IC 50 on skeletal (Nav1.4) and cardiac (Nav1.5) sodium channels is above 3000 nM. The lead molecules have the potential for future clinical development as novel therapeutics in the treatment of pain.

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Studies Examining the Relationship between the Chemical Structure of Protoxin II and Its Activity on Voltage Gated Sodium Channels

Cited by 46 publications

References 37 publications

Interaction of Tarantula Venom Peptide ProTx-II with Lipid Membranes Is a Prerequisite for Its Inhibition of Human Voltage-gated Sodium Channel NaV1.7

Interaction of Tarantula Venom Peptide ProTx-II with Lipid Membranes Is a Prerequisite for Its Inhibition of Human Voltage-gated Sodium Channel NaV1.7

Spider peptide toxin HwTx-IV engineered to bind to lipid membranes has an increased inhibitory potency at human voltage-gated sodium channel hNa V 1.7

Engineering Highly Potent and Selective Microproteins against Nav1.7 Sodium Channel for Treatment of Pain

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