Skip to main content
PMC home page
Biophysical Journal logoLink to Biophysical Journal
. 1993 Oct;65(4):1613–1619. doi: 10.1016/S0006-3495(93)81200-1

Mechanism of charybdotoxin block of a voltage-gated K+ channel.

S A Goldstein 1, C Miller 1
PMCID: PMC1225887  PMID: 7506068

Abstract

Charybdotoxin block of a Shaker K+ channel was studied in Xenopus oocyte macropatches. Toxin on rate increases linearly with toxin concentration in an ionic strength-dependent fashion and is competitively diminished by tetraethylammonium. On rate is insensitive to transmembrane voltage and to K+ on the opposite side of the membrane. Conversely, toxin off rate is insensitive to toxin concentration, ionic strength, and added tetraethylammonium but is enhanced by membrane depolarization or K+ (or Na+) in the trans solution. Charge neutralization of charybdotoxin Lys27, however, renders off rate voltage insensitive. Our results argue that block of voltage-gated K+ channels results from the binding of one toxin molecule, so that Lys27 enters the pore and interacts with K+ (or Na+) in the ion conduction pathway.

Full text

PDF
1617

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Anderson C. S., MacKinnon R., Smith C., Miller C. Charybdotoxin block of single Ca2+-activated K+ channels. Effects of channel gating, voltage, and ionic strength. J Gen Physiol. 1988 Mar;91(3):317–333. doi: 10.1085/jgp.91.3.317. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Armstrong C. M. Interaction of tetraethylammonium ion derivatives with the potassium channels of giant axons. J Gen Physiol. 1971 Oct;58(4):413–437. doi: 10.1085/jgp.58.4.413. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bontems F., Gilquin B., Roumestand C., Ménez A., Toma F. Analysis of side-chain organization on a refined model of charybdotoxin: structural and functional implications. Biochemistry. 1992 Sep 1;31(34):7756–7764. doi: 10.1021/bi00149a003. [DOI] [PubMed] [Google Scholar]
  4. Escobar L., Root M. J., MacKinnon R. Influence of protein surface charge on the bimolecular kinetics of a potassium channel peptide inhibitor. Biochemistry. 1993 Jul 13;32(27):6982–6987. doi: 10.1021/bi00078a024. [DOI] [PubMed] [Google Scholar]
  5. Giangiacomo K. M., Garcia M. L., McManus O. B. Mechanism of iberiotoxin block of the large-conductance calcium-activated potassium channel from bovine aortic smooth muscle. Biochemistry. 1992 Jul 28;31(29):6719–6727. doi: 10.1021/bi00144a011. [DOI] [PubMed] [Google Scholar]
  6. Giangiacomo K. M., Sugg E. E., Garcia-Calvo M., Leonard R. J., McManus O. B., Kaczorowski G. J., Garcia M. L. Synthetic charybdotoxin-iberiotoxin chimeric peptides define toxin binding sites on calcium-activated and voltage-dependent potassium channels. Biochemistry. 1993 Mar 9;32(9):2363–2370. doi: 10.1021/bi00060a030. [DOI] [PubMed] [Google Scholar]
  7. Gimenez-Gallego G., Navia M. A., Reuben J. P., Katz G. M., Kaczorowski G. J., Garcia M. L. Purification, sequence, and model structure of charybdotoxin, a potent selective inhibitor of calcium-activated potassium channels. Proc Natl Acad Sci U S A. 1988 May;85(10):3329–3333. doi: 10.1073/pnas.85.10.3329. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Goldstein S. A., Miller C. A point mutation in a Shaker K+ channel changes its charybdotoxin binding site from low to high affinity. Biophys J. 1992 Apr;62(1):5–7. doi: 10.1016/S0006-3495(92)81760-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hoshi T., Zagotta W. N., Aldrich R. W. Biophysical and molecular mechanisms of Shaker potassium channel inactivation. Science. 1990 Oct 26;250(4980):533–538. doi: 10.1126/science.2122519. [DOI] [PubMed] [Google Scholar]
  10. Hoshi T., Zagotta W. N., Aldrich R. W. Two types of inactivation in Shaker K+ channels: effects of alterations in the carboxy-terminal region. Neuron. 1991 Oct;7(4):547–556. doi: 10.1016/0896-6273(91)90367-9. [DOI] [PubMed] [Google Scholar]
  11. MacKinnon R., Heginbotham L., Abramson T. Mapping the receptor site for charybdotoxin, a pore-blocking potassium channel inhibitor. Neuron. 1990 Dec;5(6):767–771. doi: 10.1016/0896-6273(90)90335-d. [DOI] [PubMed] [Google Scholar]
  12. MacKinnon R., Miller C. Functional modification of a Ca2+-activated K+ channel by trimethyloxonium. Biochemistry. 1989 Oct 3;28(20):8087–8092. doi: 10.1021/bi00446a019. [DOI] [PubMed] [Google Scholar]
  13. MacKinnon R., Miller C. Mechanism of charybdotoxin block of the high-conductance, Ca2+-activated K+ channel. J Gen Physiol. 1988 Mar;91(3):335–349. doi: 10.1085/jgp.91.3.335. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. MacKinnon R., Miller C. Mutant potassium channels with altered binding of charybdotoxin, a pore-blocking peptide inhibitor. Science. 1989 Sep 22;245(4924):1382–1385. doi: 10.1126/science.2476850. [DOI] [PubMed] [Google Scholar]
  15. Miller C. Competition for block of a Ca2(+)-activated K+ channel by charybdotoxin and tetraethylammonium. Neuron. 1988 Dec;1(10):1003–1006. doi: 10.1016/0896-6273(88)90157-2. [DOI] [PubMed] [Google Scholar]
  16. Pardo L. A., Heinemann S. H., Terlau H., Ludewig U., Lorra C., Pongs O., Stühmer W. Extracellular K+ specifically modulates a rat brain K+ channel. Proc Natl Acad Sci U S A. 1992 Mar 15;89(6):2466–2470. doi: 10.1073/pnas.89.6.2466. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Park C. S., Hausdorff S. F., Miller C. Design, synthesis, and functional expression of a gene for charybdotoxin, a peptide blocker of K+ channels. Proc Natl Acad Sci U S A. 1991 Mar 15;88(6):2046–2050. doi: 10.1073/pnas.88.6.2046. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Park C. S., Miller C. Interaction of charybdotoxin with permeant ions inside the pore of a K+ channel. Neuron. 1992 Aug;9(2):307–313. doi: 10.1016/0896-6273(92)90169-e. [DOI] [PubMed] [Google Scholar]
  19. Park C. S., Miller C. Mapping function to structure in a channel-blocking peptide: electrostatic mutants of charybdotoxin. Biochemistry. 1992 Sep 1;31(34):7749–7755. doi: 10.1021/bi00149a002. [DOI] [PubMed] [Google Scholar]
  20. Schwarz T. L., Tempel B. L., Papazian D. M., Jan Y. N., Jan L. Y. Multiple potassium-channel components are produced by alternative splicing at the Shaker locus in Drosophila. Nature. 1988 Jan 14;331(6152):137–142. doi: 10.1038/331137a0. [DOI] [PubMed] [Google Scholar]
  21. Shapiro M. S., DeCoursey T. E. Permeant ion effects on the gating kinetics of the type L potassium channel in mouse lymphocytes. J Gen Physiol. 1991 Jun;97(6):1251–1278. doi: 10.1085/jgp.97.6.1251. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Stampe P., Kolmakova-Partensky L., Miller C. Mapping hydrophobic residues of the interaction surface of charybdotoxin. Biophys J. 1992 Apr;62(1):8–9. doi: 10.1016/S0006-3495(92)81761-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Sugg E. E., Garcia M. L., Reuben J. P., Patchett A. A., Kaczorowski G. J. Synthesis and structural characterization of charybdotoxin, a potent peptidyl inhibitor of the high conductance Ca2(+)-activated K+ channel. J Biol Chem. 1990 Nov 5;265(31):18745–18748. [PubMed] [Google Scholar]

Articles from Biophysical Journal are provided here courtesy of The Biophysical Society

RESOURCES