Mechanisms of sodium/calcium selectivity in sodium channels probed by cysteine mutagenesis and sulfhydryl modification

M. Teresa Pérez-García, Nipavan Chiamvimonvat, Ravi Ranjan, Jeffrey R. Balser, Gordon F. Tomaselli, Eduardo Marban

Research output: Contribution to journalArticle

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Abstract

A conserved lysine residue in the 'P loop' of domain III renders sodium channels highly selective. Conversion of this residue to glutamate, to mimic the homologous position in calcium channels, enables Ca2+ to permeate sodium channels. Because the lysine-to-glutamate mutation converts a positively charged side chain to a negative one, it has been proposed that a positive charge at this position suffices for Na+ selectivity. We tested this idea by converting the critical lysine to cysteine (K1237C) in μ1 rat skeletal sodium channels expressed in Xenopus oocytes. Selectivity of the mutant channels was then characterized before and after chemical modification to alter side-chain charge. Wild-type channels are highly selective for Na+ over Ca2+ (P(Ca)/P(Na) < 0.01). The K1237C mutation significantly increases permeability to Ca2+ (P(Ca)/P(Na) = 0.6) and Sr2+. Analogous mutations in domains I (D400C), II (E755C), and IV (A1529C) did not alter the selectivity for Na+ over Ca2+, nor did any of the domain IV mutations (G1530C, W1531C, and D1532C) that are known to affect monovalent selectivity. Interestingly, the increase in permeability to Ca2+ in K1237C cannot be reversed by simply restoring the positive charge to the side chain by using the sulfhydryl modifying reagent methanethiosulfonate ethylammonium. Single-channel studies confirmed that modified K1237C channels, which exhibit a reduced unitary conductance, remain permeable to Ca2+, with a P(Ca)/P(Na) of 0.6. We conclude that the chemical identity of the residue at position 1237 is crucial for channel selectivity. Simply rendering the 1237 side chain positive does not suffice to restore selectivity to the channel.

Original languageEnglish (US)
Pages (from-to)989-996
Number of pages8
JournalBiophysical Journal
Volume72
Issue number3
DOIs
StatePublished - Jan 1 1997
Externally publishedYes

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Sodium Channels
Mutagenesis
Cysteine
Sodium
Lysine
Calcium
Mutation
Glutamic Acid
Permeability
Sulfhydryl Reagents
Calcium Channels
Xenopus
Oocytes

ASJC Scopus subject areas

  • Biophysics

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Mechanisms of sodium/calcium selectivity in sodium channels probed by cysteine mutagenesis and sulfhydryl modification. / Pérez-García, M. Teresa; Chiamvimonvat, Nipavan; Ranjan, Ravi; Balser, Jeffrey R.; Tomaselli, Gordon F.; Marban, Eduardo.

In: Biophysical Journal, Vol. 72, No. 3, 01.01.1997, p. 989-996.

Research output: Contribution to journalArticle

Pérez-García, M. Teresa ; Chiamvimonvat, Nipavan ; Ranjan, Ravi ; Balser, Jeffrey R. ; Tomaselli, Gordon F. ; Marban, Eduardo. / Mechanisms of sodium/calcium selectivity in sodium channels probed by cysteine mutagenesis and sulfhydryl modification. In: Biophysical Journal. 1997 ; Vol. 72, No. 3. pp. 989-996.
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