Dependence of μ-conotoxin block of sodium channels on ionic strength but not on the permeating [Na+]. Implications for the distinctive mechanistic interactions between Na+ and K+ channel pore-blocking toxins and their molecular targets

Ronald A. Li, Kwokyin Hui, Robert J. French, Kazuki Sato, Charles A. Henrikson, Gordon F. Tomaselli, Eduardo Marbán

Research output: Contribution to journalArticle

Abstract

μ-Conotoxins (μ-CTXs) are Na+ channel-blocking, 22-amino acid peptides produced by the sea snail Conus geographus. Although K+ channel pore-blocking toxins show specific interactions with permeant ions and strong dependence on the ionic strength (μ), no such dependence has been reported for μ-CTX and Na+ channels. Such properties would offer insight into the binding and blocking mechanism of μ-CTX as well as functional and structural properties of the Na+ channel pore. Here we studied the effects of μ and permeant ion concentration ([Na+]) on μ-CTX block of rat skeletal muscle (μ1, Nav1.4) Na + channels. μ-CTX sensitivity of wild-type and E758Q channels increased significantly (by ∼20-fold) when μ was lowered by substituting external Na+ with equimolar sucrose (from 140 to 35 mM Na +); however, toxin block was unaltered (p > 0.05) when μ was maintained by replacement of [Na+] with N-methyl-D-glucamine (NMG+), suggesting that the enhanced sensitivity at low μ was not due to reduction in [Na+]. Single-channel recordings identified the association rate constant, kon, as the primary determinant of the changes in affinity (kon increased 40- and 333-fold for μ-CTX D2N/R13Q and D12N/R13Q, respectively, when symmetric 200 mM Na+ was reduced to 50 mM). In contrast, dissociation rates changed + channels depends critically on μ but not specifically on [Na+], contrasting with the known behavior of pore-blocking K+ channel toxins. These findings suggest that different degrees of ion interaction, underlying the fundamental conduction mechanisms of Na+ and K+ channels, are mirrored in ion interactions with pore-blocking toxins.

Original languageEnglish (US)
Pages (from-to)30912-30919
Number of pages8
JournalJournal of Biological Chemistry
Volume278
Issue number33
DOIs
StatePublished - Aug 15 2003

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Conotoxins
Sodium Channels
Ionic strength
Osmolar Concentration
Ions
Conus Snail
Oceans and Seas
Sucrose
Muscle
Rats
Structural properties
Rate constants
Skeletal Muscle
Association reactions
Amino Acids
Peptides

ASJC Scopus subject areas

  • Biochemistry

Cite this

Dependence of μ-conotoxin block of sodium channels on ionic strength but not on the permeating [Na+]. Implications for the distinctive mechanistic interactions between Na+ and K+ channel pore-blocking toxins and their molecular targets. / Li, Ronald A.; Hui, Kwokyin; French, Robert J.; Sato, Kazuki; Henrikson, Charles A.; Tomaselli, Gordon F.; Marbán, Eduardo.

In: Journal of Biological Chemistry, Vol. 278, No. 33, 15.08.2003, p. 30912-30919.

Research output: Contribution to journalArticle

Li, Ronald A. ; Hui, Kwokyin ; French, Robert J. ; Sato, Kazuki ; Henrikson, Charles A. ; Tomaselli, Gordon F. ; Marbán, Eduardo. / Dependence of μ-conotoxin block of sodium channels on ionic strength but not on the permeating [Na+]. Implications for the distinctive mechanistic interactions between Na+ and K+ channel pore-blocking toxins and their molecular targets. In: Journal of Biological Chemistry. 2003 ; Vol. 278, No. 33. pp. 30912-30919.
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T1 - Dependence of μ-conotoxin block of sodium channels on ionic strength but not on the permeating [Na+]. Implications for the distinctive mechanistic interactions between Na+ and K+ channel pore-blocking toxins and their molecular targets

AU - Li, Ronald A.

AU - Hui, Kwokyin

AU - French, Robert J.

AU - Sato, Kazuki

AU - Henrikson, Charles A.

AU - Tomaselli, Gordon F.

AU - Marbán, Eduardo

PY - 2003/8/15

Y1 - 2003/8/15

N2 - μ-Conotoxins (μ-CTXs) are Na+ channel-blocking, 22-amino acid peptides produced by the sea snail Conus geographus. Although K+ channel pore-blocking toxins show specific interactions with permeant ions and strong dependence on the ionic strength (μ), no such dependence has been reported for μ-CTX and Na+ channels. Such properties would offer insight into the binding and blocking mechanism of μ-CTX as well as functional and structural properties of the Na+ channel pore. Here we studied the effects of μ and permeant ion concentration ([Na+]) on μ-CTX block of rat skeletal muscle (μ1, Nav1.4) Na + channels. μ-CTX sensitivity of wild-type and E758Q channels increased significantly (by ∼20-fold) when μ was lowered by substituting external Na+ with equimolar sucrose (from 140 to 35 mM Na +); however, toxin block was unaltered (p > 0.05) when μ was maintained by replacement of [Na+] with N-methyl-D-glucamine (NMG+), suggesting that the enhanced sensitivity at low μ was not due to reduction in [Na+]. Single-channel recordings identified the association rate constant, kon, as the primary determinant of the changes in affinity (kon increased 40- and 333-fold for μ-CTX D2N/R13Q and D12N/R13Q, respectively, when symmetric 200 mM Na+ was reduced to 50 mM). In contrast, dissociation rates changed + channels depends critically on μ but not specifically on [Na+], contrasting with the known behavior of pore-blocking K+ channel toxins. These findings suggest that different degrees of ion interaction, underlying the fundamental conduction mechanisms of Na+ and K+ channels, are mirrored in ion interactions with pore-blocking toxins.

AB - μ-Conotoxins (μ-CTXs) are Na+ channel-blocking, 22-amino acid peptides produced by the sea snail Conus geographus. Although K+ channel pore-blocking toxins show specific interactions with permeant ions and strong dependence on the ionic strength (μ), no such dependence has been reported for μ-CTX and Na+ channels. Such properties would offer insight into the binding and blocking mechanism of μ-CTX as well as functional and structural properties of the Na+ channel pore. Here we studied the effects of μ and permeant ion concentration ([Na+]) on μ-CTX block of rat skeletal muscle (μ1, Nav1.4) Na + channels. μ-CTX sensitivity of wild-type and E758Q channels increased significantly (by ∼20-fold) when μ was lowered by substituting external Na+ with equimolar sucrose (from 140 to 35 mM Na +); however, toxin block was unaltered (p > 0.05) when μ was maintained by replacement of [Na+] with N-methyl-D-glucamine (NMG+), suggesting that the enhanced sensitivity at low μ was not due to reduction in [Na+]. Single-channel recordings identified the association rate constant, kon, as the primary determinant of the changes in affinity (kon increased 40- and 333-fold for μ-CTX D2N/R13Q and D12N/R13Q, respectively, when symmetric 200 mM Na+ was reduced to 50 mM). In contrast, dissociation rates changed + channels depends critically on μ but not specifically on [Na+], contrasting with the known behavior of pore-blocking K+ channel toxins. These findings suggest that different degrees of ion interaction, underlying the fundamental conduction mechanisms of Na+ and K+ channels, are mirrored in ion interactions with pore-blocking toxins.

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