title: Voltage-sensor conformation shapes the intra-membrane drug binding site that determines gambierol affinity in Kv channels.
abstract: Marine ladder-shaped polyether toxins are implicated in neurological symptoms of fish-borne food poisonings. The toxin gambierol, produced by the marine dinoflagellate Gambierdiscus toxicus, belongs to the group of ladder-shaped polyether toxins and inhibits Kv3.1 channels with nanomolar affinity through a mechanism of gating modification. Binding determinants for gambierol localize at the lipid-exposed interface of the pore forming S5 and S6 segments, suggesting that gambierol binds outside of the permeation pathway. To explore a possible involvement of the voltage-sensing domain (VSD), we made different chimeric channels between Kv3.1 and Kv2.1, exchanging distinct parts of the gating machinery. Our results showed that neither the electro-mechanical coupling nor the S1-S3a region of the VSD affect gambierol sensitivity. In contrast, the S3b-S4 part of the VSD (paddle motif) decreased gambierol sensitivity in Kv3.1 more than 100-fold. Structure determination by homology modeling indicated that the position of the S3b-S4 paddle and its primary structure defines the shape and∖or the accessibility of the binding site for gambierol, explaining the observed differences in gambierol affinity between the channel chimeras. Furthermore, these findings explain the observed difference in gambierol affinity for the closed and open channel configurations of Kv3.1, opening new possibilities for exploring the VSDs as selectivity determinants in drug design.
authors.affiliation: Laboratory for Molecular Biophysics, Physiology and Pharmacology, University of Antwerp, 2610, Antwerp, Belgium.
authors.forename: Ivan
authors.initials: I
authors.lastname: Kopljar
authors.affiliation: CINECA, Sede di Roma, Via dei Tizii 6, 00185, Rome, Italy.
authors.forename: Alessandro
authors.initials: A
authors.lastname: Grottesi
authors.affiliation: Laboratory for Molecular Biophysics, Physiology and Pharmacology, University of Antwerp, 2610, Antwerp, Belgium.
authors.forename: Tessa
authors.initials: T
authors.lastname: de Block
authors.affiliation: Department of Chemistry, University of Utah, Salt Lake City, UT, 84112-0850, USA.
authors.forename: Jon D
authors.initials: JD
authors.lastname: Rainier
authors.affiliation: Toxicology and Pharmacology, University of Leuven Campus Gasthuisberg, 3000, Leuven, Belgium.
authors.forename: Jan
authors.initials: J
authors.lastname: Tytgat
authors.affiliation: Laboratory for Molecular Biophysics, Physiology and Pharmacology, University of Antwerp, 2610, Antwerp, Belgium.
authors.forename: Alain J
authors.initials: AJ
authors.lastname: Labro
authors.affiliation: Laboratory for Molecular Biophysics, Physiology and Pharmacology, University of Antwerp, 2610, Antwerp, Belgium. Electronic address: dirk.snyders@uantwerpen.be.
authors.forename: Dirk J
authors.initials: DJ
authors.lastname: Snyders
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D002922:Ciguatoxins / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D051379:Mice; D008958:Models, Molecular; D050506:Mutant Chimeric Proteins; D018408:Patch-Clamp Techniques; D026902:Potassium Channel Blockers / Q000494:pharmacology*; D011487:Protein Conformation; D051669:Shab Potassium Channels / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism*; D051670:Shaw Potassium Channels / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism*; D035964:Oxocins
chemical_list: D002922:Ciguatoxins; D050506:Mutant Chimeric Proteins; D026902:Potassium Channel Blockers; D051669:Shab Potassium Channels; D051670:Shaw Potassium Channels; C435770:gambierol; D035964:Oxocins
keywords: Ciguatoxins; Electrophysiology; Gating modifier; Polycyclic ether toxin; Voltage-gated potassium channel