Chemical Synthesis, Proper Folding, Na<sub>v</sub> Channel Selectivity Profile and Analgesic Properties of the Spider Peptide Phlotoxin 1
Phlotoxin-1 (PhlTx1) is a peptide previously identified in tarantula venom (<i>Phlogius</i> species) that belongs to the inhibitory cysteine-knot (ICK) toxin family. Like many ICK-based spider toxins, the synthesis of PhlTx1 appears particularly challenging, mostly for obtaining appropri...
| Main Authors: | , , , , , , , , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
MDPI AG
2019-06-01
|
| Series: | Toxins |
| Subjects: | |
| Online Access: | https://www.mdpi.com/2072-6651/11/6/367 |
| _version_ | 1798000007139295232 |
|---|---|
| author | Sébastien Nicolas Claude Zoukimian Frank Bosmans Jérôme Montnach Sylvie Diochot Eva Cuypers Stephan De Waard Rémy Béroud Dietrich Mebs David Craik Didier Boturyn Michel Lazdunski Jan Tytgat Michel De Waard |
| author_facet | Sébastien Nicolas Claude Zoukimian Frank Bosmans Jérôme Montnach Sylvie Diochot Eva Cuypers Stephan De Waard Rémy Béroud Dietrich Mebs David Craik Didier Boturyn Michel Lazdunski Jan Tytgat Michel De Waard |
| author_sort | Sébastien Nicolas |
| collection | DOAJ |
| description | Phlotoxin-1 (PhlTx1) is a peptide previously identified in tarantula venom (<i>Phlogius</i> species) that belongs to the inhibitory cysteine-knot (ICK) toxin family. Like many ICK-based spider toxins, the synthesis of PhlTx1 appears particularly challenging, mostly for obtaining appropriate folding and concomitant suitable disulfide bridge formation. Herein, we describe a procedure for the chemical synthesis and the directed sequential disulfide bridge formation of PhlTx1 that allows for a straightforward production of this challenging peptide. We also performed extensive functional testing of PhlTx1 on 31 ion channel types and identified the voltage-gated sodium (Na<sub>v</sub>) channel Na<sub>v</sub>1.7 as the main target of this toxin. Moreover, we compared PhlTx1 activity to 10 other spider toxin activities on an automated patch-clamp system with Chinese Hamster Ovary (CHO) cells expressing human Na<sub>v</sub>1.7. Performing these analyses in reproducible conditions allowed for classification according to the potency of the best natural Na<sub>v</sub>1.7 peptide blockers. Finally, subsequent in vivo testing revealed that intrathecal injection of PhlTx1 reduces the response of mice to formalin in both the acute pain and inflammation phase without signs of neurotoxicity. PhlTx1 is thus an interesting toxin to investigate Na<sub>v</sub>1.7 involvement in cellular excitability and pain. |
| first_indexed | 2024-04-11T11:13:25Z |
| format | Article |
| id | doaj.art-e86d22f9474449df85156f8c0d4cf8f5 |
| institution | Directory Open Access Journal |
| issn | 2072-6651 |
| language | English |
| last_indexed | 2024-04-11T11:13:25Z |
| publishDate | 2019-06-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Toxins |
| spelling | doaj.art-e86d22f9474449df85156f8c0d4cf8f52022-12-22T04:27:19ZengMDPI AGToxins2072-66512019-06-0111636710.3390/toxins11060367toxins11060367Chemical Synthesis, Proper Folding, Na<sub>v</sub> Channel Selectivity Profile and Analgesic Properties of the Spider Peptide Phlotoxin 1Sébastien Nicolas0Claude Zoukimian1Frank Bosmans2Jérôme Montnach3Sylvie Diochot4Eva Cuypers5Stephan De Waard6Rémy Béroud7Dietrich Mebs8David Craik9Didier Boturyn10Michel Lazdunski11Jan Tytgat12Michel De Waard13Institut du Thorax, Inserm UMR 1087/CNRS UMR 6291, LabEx “Ion Channels, Science & Therapeutics”, F-44007 Nantes, FranceSmartox Biotechnology, 6 rue des Platanes, F-38120 Saint-Egrève, FranceFaculty of Medicine and Health Sciences, Department of Basic and Applied Medical Sciences, 9000 Gent, BelgiumInstitut du Thorax, Inserm UMR 1087/CNRS UMR 6291, LabEx “Ion Channels, Science & Therapeutics”, F-44007 Nantes, FranceUniversité Côte d’Azur, CNRS UMR7275, Institut de Pharmacologie Moléculaire et Cellulaire, 660 route des lucioles, 6560 Valbonne, FranceToxicology and Pharmacology, University of Leuven, Campus Gasthuisberg, P.O. Box 922, Herestraat 49, 3000 Leuven, BelgiumInstitut du Thorax, Inserm UMR 1087/CNRS UMR 6291, LabEx “Ion Channels, Science & Therapeutics”, F-44007 Nantes, FranceSmartox Biotechnology, 6 rue des Platanes, F-38120 Saint-Egrève, FranceInstitute of Legal Medicine, University of Frankfurt, Kennedyallee 104, 60488 Frankfurt, GermanyInstitute for Molecular Bioscience, University of Queensland, Brisbane 4072, AustraliaDepartment of Molecular Chemistry, Univ. Grenoble Alpes, CNRS, 570 rue de la chimie, CS 40700, 38000 Grenoble, FranceUniversité Côte d’Azur, CNRS UMR7275, Institut de Pharmacologie Moléculaire et Cellulaire, 660 route des lucioles, 6560 Valbonne, FranceToxicology and Pharmacology, University of Leuven, Campus Gasthuisberg, P.O. Box 922, Herestraat 49, 3000 Leuven, BelgiumInstitut du Thorax, Inserm UMR 1087/CNRS UMR 6291, LabEx “Ion Channels, Science & Therapeutics”, F-44007 Nantes, FrancePhlotoxin-1 (PhlTx1) is a peptide previously identified in tarantula venom (<i>Phlogius</i> species) that belongs to the inhibitory cysteine-knot (ICK) toxin family. Like many ICK-based spider toxins, the synthesis of PhlTx1 appears particularly challenging, mostly for obtaining appropriate folding and concomitant suitable disulfide bridge formation. Herein, we describe a procedure for the chemical synthesis and the directed sequential disulfide bridge formation of PhlTx1 that allows for a straightforward production of this challenging peptide. We also performed extensive functional testing of PhlTx1 on 31 ion channel types and identified the voltage-gated sodium (Na<sub>v</sub>) channel Na<sub>v</sub>1.7 as the main target of this toxin. Moreover, we compared PhlTx1 activity to 10 other spider toxin activities on an automated patch-clamp system with Chinese Hamster Ovary (CHO) cells expressing human Na<sub>v</sub>1.7. Performing these analyses in reproducible conditions allowed for classification according to the potency of the best natural Na<sub>v</sub>1.7 peptide blockers. Finally, subsequent in vivo testing revealed that intrathecal injection of PhlTx1 reduces the response of mice to formalin in both the acute pain and inflammation phase without signs of neurotoxicity. PhlTx1 is thus an interesting toxin to investigate Na<sub>v</sub>1.7 involvement in cellular excitability and pain.https://www.mdpi.com/2072-6651/11/6/367spider toxindirected disulfide bond formationNa<sub>v</sub> channel activityNa<sub>v</sub>1.7pain targetautomated patch-clamp |
| spellingShingle | Sébastien Nicolas Claude Zoukimian Frank Bosmans Jérôme Montnach Sylvie Diochot Eva Cuypers Stephan De Waard Rémy Béroud Dietrich Mebs David Craik Didier Boturyn Michel Lazdunski Jan Tytgat Michel De Waard Chemical Synthesis, Proper Folding, Na<sub>v</sub> Channel Selectivity Profile and Analgesic Properties of the Spider Peptide Phlotoxin 1 Toxins spider toxin directed disulfide bond formation Na<sub>v</sub> channel activity Na<sub>v</sub>1.7 pain target automated patch-clamp |
| title | Chemical Synthesis, Proper Folding, Na<sub>v</sub> Channel Selectivity Profile and Analgesic Properties of the Spider Peptide Phlotoxin 1 |
| title_full | Chemical Synthesis, Proper Folding, Na<sub>v</sub> Channel Selectivity Profile and Analgesic Properties of the Spider Peptide Phlotoxin 1 |
| title_fullStr | Chemical Synthesis, Proper Folding, Na<sub>v</sub> Channel Selectivity Profile and Analgesic Properties of the Spider Peptide Phlotoxin 1 |
| title_full_unstemmed | Chemical Synthesis, Proper Folding, Na<sub>v</sub> Channel Selectivity Profile and Analgesic Properties of the Spider Peptide Phlotoxin 1 |
| title_short | Chemical Synthesis, Proper Folding, Na<sub>v</sub> Channel Selectivity Profile and Analgesic Properties of the Spider Peptide Phlotoxin 1 |
| title_sort | chemical synthesis proper folding na sub v sub channel selectivity profile and analgesic properties of the spider peptide phlotoxin 1 |
| topic | spider toxin directed disulfide bond formation Na<sub>v</sub> channel activity Na<sub>v</sub>1.7 pain target automated patch-clamp |
| url | https://www.mdpi.com/2072-6651/11/6/367 |
| work_keys_str_mv | AT sebastiennicolas chemicalsynthesisproperfoldingnasubvsubchannelselectivityprofileandanalgesicpropertiesofthespiderpeptidephlotoxin1 AT claudezoukimian chemicalsynthesisproperfoldingnasubvsubchannelselectivityprofileandanalgesicpropertiesofthespiderpeptidephlotoxin1 AT frankbosmans chemicalsynthesisproperfoldingnasubvsubchannelselectivityprofileandanalgesicpropertiesofthespiderpeptidephlotoxin1 AT jeromemontnach chemicalsynthesisproperfoldingnasubvsubchannelselectivityprofileandanalgesicpropertiesofthespiderpeptidephlotoxin1 AT sylviediochot chemicalsynthesisproperfoldingnasubvsubchannelselectivityprofileandanalgesicpropertiesofthespiderpeptidephlotoxin1 AT evacuypers chemicalsynthesisproperfoldingnasubvsubchannelselectivityprofileandanalgesicpropertiesofthespiderpeptidephlotoxin1 AT stephandewaard chemicalsynthesisproperfoldingnasubvsubchannelselectivityprofileandanalgesicpropertiesofthespiderpeptidephlotoxin1 AT remyberoud chemicalsynthesisproperfoldingnasubvsubchannelselectivityprofileandanalgesicpropertiesofthespiderpeptidephlotoxin1 AT dietrichmebs chemicalsynthesisproperfoldingnasubvsubchannelselectivityprofileandanalgesicpropertiesofthespiderpeptidephlotoxin1 AT davidcraik chemicalsynthesisproperfoldingnasubvsubchannelselectivityprofileandanalgesicpropertiesofthespiderpeptidephlotoxin1 AT didierboturyn chemicalsynthesisproperfoldingnasubvsubchannelselectivityprofileandanalgesicpropertiesofthespiderpeptidephlotoxin1 AT michellazdunski chemicalsynthesisproperfoldingnasubvsubchannelselectivityprofileandanalgesicpropertiesofthespiderpeptidephlotoxin1 AT jantytgat chemicalsynthesisproperfoldingnasubvsubchannelselectivityprofileandanalgesicpropertiesofthespiderpeptidephlotoxin1 AT micheldewaard chemicalsynthesisproperfoldingnasubvsubchannelselectivityprofileandanalgesicpropertiesofthespiderpeptidephlotoxin1 |