Specificity for block by saxitoxin and divalent cations at a residue which determines sensitivity of sodium channel subtypes to guanidinium toxins

Détails

ID Serval
serval:BIB_32160F136872
Type
Article: article d'un périodique ou d'un magazine.
Collection
Publications
Institution
Titre
Specificity for block by saxitoxin and divalent cations at a residue which determines sensitivity of sodium channel subtypes to guanidinium toxins
Périodique
Journal of General Physiology
Auteur(s)
Favre  I., Moczydlowski  E., Schild  L.
ISSN
0022-1295 (Print)
Statut éditorial
Publié
Date de publication
08/1995
Volume
106
Numéro
2
Pages
203-29
Notes
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S. --- Old month value: Aug
Résumé
bTyrosine 401 of the skeletal muscle isoform (mu 1) of the rat muscle Na channel is an important determinant of high affinity block by tetrodotoxin (TTX) and saxitoxin (STX) in Na-channel isoforms. In mammalian heart Na channels, this residue is substituted by cysteine, which results in low affinity for TTX/STX and enhanced sensitivity to block by Zn2+ and Cd2+. In this study, we investigated the molecular basis for high affinity block of Na channels by STX and divalent cations by measuring inhibition of macroscopic Na+ current for a series of point mutations at residue Tyr401 of the rat mu 1 Na channel expressed in Xenopus oocytes. Substitution of Tyr401 by Gly, Ala, Ser, Cys, Asp, His, Trp, and Phe produced functional Na+ currents without major perturbation of gating or ionic selectivity. High affinity block by STX and neosaxitoxin (NEO) with Ki values in the range of 2.6-18 nM required Tyr, Phe, or Trp, suggestive of an interaction between an aromatic ring and a guanidinium group of the toxin. The Cys mutation resulted in a 7- and 23-fold enhancement of the dissociation rate of STX and NEO, respectively, corresponding to rapid toxin dissociation rates of cardiac Na channels. High affinity block by Zn2+ (Ki = 8-23 microM) required Cys, His, or Asp, three residues commonly found to coordinate directly with Zn2+ in metalloproteins. For the Cys mutant of mu 1 and also for the cardiac isoform Na channel (rh1) expressed in the L6 rat muscle cell line, inhibition of macroscopic Na+ conductance by Zn2+ reached a plateau at 85-90% inhibition, suggesting the presence of a substate current. The Asp mutant also displayed enhanced affinity for inhibition of conductance by Ca2+ (Ki = 0.3 mM vs approximately 40 mM in wild type), but block by Ca2+ was incomplete, saturating at approximately 69% inhibition. In contrast, Cd2+ completely blocked macroscopic current in the Cys mutant and the L6 cell line. These results imply that the magnitude of substate current depends on the particular residue at position 401 and the species of divalent cation. The His mutant also exhibited enhanced sensitivity to block by H+ with a pKa of approximately 7.5 for the His imidazole group. Our findings provide further evidence that residue 401 of mu 1 is located within the outer vestibule of the Na channel but external to the single-filing region for permeant ions.
Mots-clé
Animals Calcium/metabolism Cell Line Cell Membrane Permeability/*drug effects/physiology Female Ion Channel Gating/*drug effects/physiology Muscle, Skeletal/cytology/*metabolism Neuromuscular Blocking Agents/pharmacology Oocytes/metabolism Patch-Clamp Techniques Point Mutation Rats Saxitoxin/analogs & derivatives/*pharmacology Sodium/*metabolism *Sodium Channel Blockers Tyrosine/genetics/metabolism Xenopus laevis
Pubmed
Web of science
Open Access
Oui
Création de la notice
24/01/2008 13:56
Dernière modification de la notice
20/08/2019 14:17
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