A membrane-access mechanism of ion channel inhibition by voltage sensor toxins from spider venom


Venomous animals produce small protein toxins that inhibit ion channels with high affinity. In several well-studied cases the inhibitory proteins are water-soluble and bind at a channel's aqueous-exposed extracellular surface1,2,3,4. Here we show that a voltage-sensor toxin (VSTX1) from the Chilean Rose Tarantula (Grammostola spatulata) reaches its target by partitioning into the lipid membrane. Lipid membrane partitioning serves two purposes: to localize the toxin in the membrane where the voltage sensor resides and to exploit the free energy of partitioning to achieve apparent high-affinity inhibition. VSTX1, small hydrophobic poisons and anaesthetic molecules reveal a common theme of voltage sensor inhibition through lipid membrane access. The apparent requirement for such access is consistent with the recent proposal that the sensor in voltage-dependent K+ channels is located at the membrane–protein interface5,6.

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Figure 1: VSTX1 binds to the KvAP channel with high affinity in lipid membranes and low affinity in detergent micelles.
Figure 2: VSTX1 binds to lipid membranes.
Figure 3: Increase in tryptophan fluorescence of VSTX1 on partitioning into membranes.
Figure 4: Hydrophobic face of voltage sensor toxins is conserved.


  1. 1

    Miller, C., Moczydlowski, E., Latorre, R. & Phillips, M. Charybdotoxin, a protein inhibitor of single Ca2+-activated K+ channels from mammalian skeletal muscle. Nature 313, 316–318 (1985)

    ADS  CAS  Article  Google Scholar 

  2. 2

    Hidalgo, P. & MacKinnon, R. Revealing the architecture of a K+ channel pore through mutant cycles with a peptide inhibitor. Science 268, 307–310 (1995)

    ADS  CAS  Article  Google Scholar 

  3. 3

    Imredy, J. P., Chen, C. & MacKinnon, R. A snake toxin inhibitor of inward rectifier potassium channel ROMK1. Biochemistry 37, 14867–14874 (1998)

    CAS  Article  Google Scholar 

  4. 4

    Hugues, M., Romey, G., Duval, D., Vincent, J. P. & Lazdunski, M. Apamin as a selective blocker of the calcium-dependent potassium channel in neuroblastoma cells: voltage-clamp and biochemical characterization of the toxin receptor. Proc. Natl Acad. Sci. USA 79, 1308–1312 (1982)

    ADS  CAS  Article  Google Scholar 

  5. 5

    Jiang, Y., Ruta, V., Chen, J., Lee, A. & MacKinnon, R. The principle of gating charge movement in a voltage-dependent K+ channel. Nature 423, 42–48 (2003)

    ADS  CAS  Article  Google Scholar 

  6. 6

    Jiang, Y. et al. X-ray structure of a voltage-dependent K+ channel. Nature 423, 33–41 (2003)

    ADS  CAS  Article  Google Scholar 

  7. 7

    Miller, C. Diffusion-controlled binding of a peptide neurotoxin to its K+ channel receptor. Biochemistry 29, 5320–5325 (1990)

    CAS  Article  Google Scholar 

  8. 8

    Escobar, L., Root, M. J. & MacKinnon, R. Influence of protein surface charge on the bimolecular kinetics of a potassium channel peptide inhibitor. Biochemistry 32, 6982–6987 (1993)

    CAS  Article  Google Scholar 

  9. 9

    Goldstein, S. A., Pheasant, D. J. & Miller, C. The charybdotoxin receptor of a Shaker K+ channel: peptide and channel residues mediating molecular recognition. Neuron 12, 1377–1388 (1994)

    CAS  Article  Google Scholar 

  10. 10

    Ranganathan, R., Lewis, J. H. & MacKinnon, R. Spatial localization of the K+ channel selectivity filter by mutant cycle-based structure analysis. Neuron 16, 131–139 (1996)

    CAS  Article  Google Scholar 

  11. 11

    Swartz, K. J. & MacKinnon, R. Hanatoxin modifies the gating of a voltage-dependent K+ channel through multiple binding sites. Neuron 18, 665–673 (1997)

    CAS  Article  Google Scholar 

  12. 12

    Swartz, K. J. & MacKinnon, R. Mapping the receptor site for hanatoxin, a gating modifier of voltage-dependent K+ channels. Neuron 18, 675–682 (1997)

    CAS  Article  Google Scholar 

  13. 13

    Swartz, K. J. & MacKinnon, R. An inhibitor of the Kv2.1 potassium channel isolated from the venom of a Chilean tarantula. Neuron 15, 941–949 (1995)

    CAS  Article  Google Scholar 

  14. 14

    Ruta, V., Jiang, Y., Lee, A., Chen, J. & MacKinnon, R. Functional analysis of an archeabacterial voltage-dependent K+ channel. Nature 422, 180–185 (2003)

    ADS  CAS  Article  Google Scholar 

  15. 15

    Winterfield, J. R. & Swartz, K. J. A hot spot for the interaction of gating modifier toxins with voltage-dependent ion channels. J. Gen. Physiol. 116, 637–644 (2000)

    CAS  Article  Google Scholar 

  16. 16

    Li-Smerin, Y. & Swartz, K. J. Gating modifier toxins reveal a conserved structural motif in voltage- gated Ca2+ and K+ channels. Proc. Natl Acad. Sci. USA 95, 8585–8589 (1998)

    ADS  CAS  Article  Google Scholar 

  17. 17

    Lee, C. W. et al. Solution Structure and Functional Characterization of SGTx1, a Modifier of Kv2.1 Channel Gating. Biochemistry 43, 890–897 (2004)

    CAS  Article  Google Scholar 

  18. 18

    Takahashi, H. et al. Solution structure of hanatoxin1, a gating modifier of voltage-dependent K(+ ) channels: common surface features of gating modifier toxins. J. Mol. Biol. 297, 771–780 (2000)

    CAS  Article  Google Scholar 

  19. 19

    Manoleras, N. & Norton, R. S. Three-dimensional structure in solution of neurotoxin III from the sea anemone Anemonia sulcata. Biochemistry 33, 11051–11061 (1994)

    CAS  Article  Google Scholar 

  20. 20

    Wang, J. et al. Functional mapping of the molecular surface of a gating modifier toxin for voltage-gated K+ channels. Biophys. J. 84, 218a (2003)

    Google Scholar 

  21. 21

    Lee, H. C., Wang, J. M. & Swartz, K. J. Interaction between extracellular Hanatoxin and the resting conformation of the voltage-sensor paddle in Kv channels. Neuron 40, 527–536 (2003)

    CAS  Article  Google Scholar 

  22. 22

    Jiang, Q. X. Spherical reconstruction: a novel method for structure determination from cryo-EM images of membrane proteins in small vesicles. Thesis, Yale Univ. (2001)

    Google Scholar 

  23. 23

    Garcia, M. L., Garcia-Calvo, M., Hidalgo, P., Lee, A. & MacKinnon, R. Purification and characterization of three inhibitors of voltage-dependent K+ channels from Leiurus quinquestriatus var. hebraeus venom. Biochemistry 33, 6834–6839 (1994)

    CAS  Article  Google Scholar 

  24. 24

    Ladokhin, A. S., Jayasinghe, S. & White, S. H. How to measure and analyze tryptophan fluorescence in membranes properly, and why bother? Anal. Biochem. 285, 235–245 (2000)

    CAS  Article  Google Scholar 

  25. 25

    Almeida, P. F. F. & Vaz, W. L. C. in Handbook of Biological Physics (eds Lipowsky, R. & Sackmann, E.) 305–357 (Elsevier Science, Netherlands, 1995)

    Google Scholar 

  26. 26

    Schurr, J. M. The role of diffusion in enzyme kinetics. Biophys. J. 10, 717–727 (1970)

    ADS  CAS  Article  Google Scholar 

  27. 27

    Hille, B. Ion Channels of Excitable Membranes 635–662 (Sinauer Associates, Sunderland, MA, 2001)

    Google Scholar 

  28. 28

    Heginbotham, L., LeMasurier, M., Kolmakova-Partensky, L. & Miller, C. Single streptomyces lividans K(+ ) channels: functional asymmetries and sidedness of proton activation. J. Gen. Physiol. 114, 551–560 (1999)

    CAS  Article  Google Scholar 

  29. 29

    Diochot, S., Drici, M. D., Moinier, D., Fink, M. & Lazdunski, M. Effects of phrixotoxins on the Kv4 family of potassium channels and implications for the role of Ito1 in cardiac electrogenesis. Br. J. Pharmacol. 126, 251–263 (1999)

    CAS  Article  Google Scholar 

  30. 30

    Escoubas, P., Diochot, S., Celerier, M. L., Nakajima, T. & Lazdunski, M. Novel tarantula toxins for subtypes of voltage-dependent potassium channels in the Kv2 and Kv4 subfamilies. Mol. Pharmacol. 62, 48–57 (2002)

    CAS  Article  Google Scholar 

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We thank D. King for toxin synthesis for initial experiments, F. Valiyaveetil for help with fluorescence experiments, V. Ruta for help with toxin purification and electrophysiology and S. Long for providing AgTx2. This work was supported by a National Institutes of Health grant to R.M. R.M. is an investigator in the Howard Hughes Medical Institute.

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Correspondence to Roderick MacKinnon.

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Lee, S., MacKinnon, R. A membrane-access mechanism of ion channel inhibition by voltage sensor toxins from spider venom. Nature 430, 232–235 (2004). https://doi.org/10.1038/nature02632

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