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Unformatted text preview: Tarantula Huwentoxin-IV Inhibits Neuronal Sodium Channels by Binding to Receptor Site 4 and Trapping the Domain II Voltage Sensor in the Closed Configuration * □ S Received for publication, October 11, 2007, and in revised form, June 11, 2008 Published, JBC Papers in Press, July 14, 2008, DOI 10.1074/jbc.M708447200 Yucheng Xiao ‡§ , Jon-Paul Bingham ¶ , Weiguo Zhu ‡ , Edward Moczydlowski , Songping Liang §1 , and Theodore R. Cummins ‡2 From the ‡ Department of Pharmacology and Toxicology, Stark Neurosciences Research Institute, Indiana University School of Medicine, Indianapolis, Indiana 46202, ¶ Department of Molecular Biosciences and Bioengineering, University of Hawaii, Honolulu, Hawaii 96822, Department of Biology, Clarkson University, Potsdam, New York 13699, and § Life Sciences College, Hunan Normal University, Changsha, Hunan 410081, China Peptide toxins with high affinity, divergent pharmacological functions, and isoform-specific selectivity are powerful tools for investigating the structure-function relationships of voltage- gated sodium channels (VGSCs). Although a number of inter- esting inhibitors have been reported from tarantula venoms, lit- tle is known about the mechanism for their interaction with VGSCs.Weshowthathuwentoxin-IV(HWTX-IV),a35-residue peptide from tarantula Ornithoctonus huwena venom, preferen- tially inhibits neuronal VGSC subtypes rNav1.2, rNav1.3, and hNav1.7 compared with muscle subtypes rNav1.4 and hNav1.5. Of the five VGSCs examined, hNav1.7 was most sensitive to HWTX-IV (IC 50 26 n M ). Following application of 1 M HWTX-IV, hNav1.7 currents could only be elicited with extreme depolarizations ( > 100 mV). Recovery of hNav1.7 channels from HWTX-IV inhibition could be induced by extreme depolarizations or moderate depolarizations lasting several minutes. Site-directed mutagenesis analysis indicated that the toxin docked at neurotoxin receptor site 4 located at the extracellular S3-S4 linker of domain II. Mutations E818Q and D816N in hNav1.7 decreased toxin affinity for hNav1.7 by 300-fold, whereas the reverse mutations in rNav1.4 (N655D/ Q657E) and the corresponding mutations in hNav1.5 (R812D/ S814E) greatly increased the sensitivity of the muscle VGSCs to HWTX-IV. Our data identify a novel mechanism for sodium channel inhibition by tarantula toxins involving binding to neu- rotoxinreceptorsite4.Incontrasttoscorpion-toxinsthattrap the IIS4 voltage sensor in an outward configuration, we propose that HWTX-IV traps the voltage sensor of domain II in the inward, closed configuration. Voltage-gated sodium channels (VGSCs) 3 are important transmembrane proteins expressed in most excitable tissues. The opening of the pore-forming subunit is responsible for the rapid depolarizing phase of action potentials. Nine dis- tinct VGSC subunit subtypes (Nav1.1–1.9) have been cloned from mammals (1, 2). The subunits share over 75% sequence similarity with each other and often exhibit similar functional properties. Because of these similarities, biologi-functional properties....
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This note was uploaded on 01/27/2012 for the course ECOLOGY 300 taught by Professor Zumdahli during the Spring '11 term at St. Mary NE.
- Spring '11