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Purification of the tetrodotoxin-binding component associated with the voltage-sensitive sodium channel from Electrophorus electricus electroplax membranes

Agnew, William S. and Levinson, Simon R. and Brabson, John S. and Raftery, Michael A. (1978) Purification of the tetrodotoxin-binding component associated with the voltage-sensitive sodium channel from Electrophorus electricus electroplax membranes. Proceedings of the National Academy of Sciences of the United States of America, 75 (6). pp. 2606-2610. ISSN 0027-8424. https://resolver.caltech.edu/CaltechAUTHORS:20120724-152628705

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Abstract

The tetrodotoxin-binding component associated with the voltage-sensitive sodium channel from electroplax membranes of Electrophorus electricus has been purified. The toxin-binding site could be efficiently solubilized with Lubrol-PX, resulting in an extract of high initial specific activity. Purification was facilitated by the development of a rapid, quantitative binding assay. The binding component was stabilized during purification by the use of mixed lipid/detergent micelles of defined composition, and by the saturation of the site with tetrodotoxin. The purification was achieved by means of a highly selective adsorption of the toxin-binding component to DEASE-Sephadex A-25, followed by desorption at high ionic strength and chromatography over Sepharose 6B. Final peak specific activities were at least 50% of the specific activity expected for a pure, undenatured toxin-binding componenet of 230,000 molecular weight. The purified material exhibited a sedimentation coefficient of approximately 8 S and an unusual Stokes radius of 95 A. Purified material showed a relatively simple pattern on sodium dodecyl sulfate/polyacrylamide gel electrophoresis, being comprised of only three polypeptides.


Item Type:Article
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http://www.pnas.org/content/75/6/2606.abstractPublisherUNSPECIFIED
Additional Information:© 1978 National Academy of Sciences. Communicated by Harden M. McConnell, March 6, 1978. The authors wish to thank Dr. T. Benzer and Mr. F. Stackhouse for initial development of TTX-binding assays and Mrs. Valerie Purvis for preparation of technical illustrations and manuscript typing. This work was supported by U.S. Public Health Service Grant NS 12018, by a grant from the Sloan Foundation, by National Institutes of Health Postdoctoral Fellowships to W.S.A., S.R.L., and J.S.B., and a Fellowship from the Muscular Dystrophy Association of America to W.S.A. 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. §1734 solely to indicate this fact.
Funders:
Funding AgencyGrant Number
Public Health ServiceNS 12018
Sloan FoundationUNSPECIFIED
NIH Postdoctoral FellowshipUNSPECIFIED
Muscular Dystrophy Association of America FellowshipUNSPECIFIED
Subject Keywords:receptor, saxitoxin, electrical excitability, mixed micelle
Issue or Number:6
Record Number:CaltechAUTHORS:20120724-152628705
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20120724-152628705
Official Citation: W S Agnew, S R Levinson, J S Brabson, and M A Raftery Purification of the tetrodotoxin-binding component associated with the voltage-sensitive sodium channel from Electrophorus electricus electroplax membranes PNAS 1978 75 (6) 2606-2610
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:32692
Collection:CaltechAUTHORS
Deposited By: Ruth Sustaita
Deposited On:24 Jul 2012 23:01
Last Modified:03 Oct 2019 04:03

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