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A Cation-π Interaction Discriminates among Sodium Channels That Are Either Sensitive or Resistant to Tetrodotoxin Block

Santarelli, Vincent P. and Eastwood, Amy L. and Dougherty, Dennis A. and Horn, Richard and Ahern, Christopher A. (2007) A Cation-π Interaction Discriminates among Sodium Channels That Are Either Sensitive or Resistant to Tetrodotoxin Block. Journal of Biological Chemistry, 282 (11). pp. 8044-8051. ISSN 0021-9258. doi:10.1074/jbc.M611334200.

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Voltage-gated sodium channels control the upstroke of the action potential in excitable cells of nerve and muscle tissue, making them ideal targets for exogenous toxins that aim to squelch electrical excitability. One such toxin, tetrodotoxin (TTX), blocks sodium channels with nanomolar affinity only when an aromatic Phe or Tyr residue is present at a specific location in the external vestibule of the ion-conducting pore. To test whether TTX is attracted to Tyr401 of NaV1.4 through a cation-{pi} interaction, this aromatic residue was replaced with fluorinated derivatives of Phe using in vivo nonsense suppression. Consistent with a cation-{pi} interaction, increased fluorination of Phe401, which reduces the negative electrostatic potential on the aromatic face, caused a monotonic increase in the inhibitory constant for block. Trifluorination of the aromatic ring decreased TTX affinity by ~50-fold, a reduction similar to that caused by replacement with the comparably hydrophobic residue Leu. Furthermore, we show that an energetically equivalent cation-{pi} interaction underlies both use-dependent and tonic block by TTX. Our results are supported by high level ab initio quantum mechanical calculations applied to a model of TTX binding to benzene. Our analysis suggests that the aromatic side chain faces the permeation pathway where it orients TTX optimally and interacts with permeant ions. These results are the first of their kind to show the incorporation of unnatural amino acids into a voltage-gated sodium channel and demonstrate that a cation-{pi} interaction is responsible for the obligate nature of an aromatic at this position in TTX-sensitive sodium channels.

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Dougherty, Dennis A.0000-0003-1464-2461
Additional Information:© 2007 the American Society for Biochemistry and Molecular Biology. Received for publication, December 11, 2006 , and in revised form, January 11, 2007. Originally published In Press as doi:10.1074/jbc.M611334200 on January 19, 2007. We thank Mary Y. Ryan for help with oocytes and molecular biology. We also thank Drs. Lipkind and Fozzard for helpful discussions about TTX block. This work was supported by Grants GM079427 and NS34407 from the National Institutes of Health. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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Issue or Number:11
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ID Code:7625
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Deposited On:14 Mar 2007
Last Modified:08 Nov 2021 20:44

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