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Identification of the polypeptide chains in Torpedo californica electroplax membranes that interact with a local anesthetic analog

Blanchard, Steven G. and Raftery, Michael A. (1979) Identification of the polypeptide chains in Torpedo californica electroplax membranes that interact with a local anesthetic analog. Proceedings of the National Academy of Sciences of the United States of America, 76 (1). pp. 81-85. ISSN 0027-8424.

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Procaine amide azide, a derivative of the local anesthetic procaine amide, was prepared and its interactions with acetylcholine receptor-rich membrane fragments from Torpedo californica electroplax were studied. Procaine amide azide was radiolabeled by a method that may be of general use for the preparation of other radioactive tertiary amines. When low concentrations of 3H-labeled procaine amide azide were photolyzed in the presence of receptor-rich membrane preparations, a simple pattern of incorporated radioactivity was seen after electrophoresis on sodium dodecyl sulfate/polyacrylamide gels. Only two major labeled bands were seen, corresponding to apparent Mr of about 43,000 and 90,000. When the length of the gels was increased, the labeled band of lower Mr was resolved into two labeled proteins, one major and one minor, with apparent Mr of 43,000 and 40,000, respectively. The radioactivity incorporated into the protein of Mr 40,000 could be attributed to interaction of [3H]procaine amide azide with cholinergic ligand-binding sites, whereas labeling of the polypeptide of Mr 43,000 appears to represent interaction of the photolabile derivative with another class of sites. The labeling component of 90,000 Mr could be removed by preparation of membrane fragments in iodoacetamide-containing buffer and therefore appeared unrelated to the acetylcholine receptor.

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Additional Information:© 1979 by the National Academy of Sciences. Communicated by James Bonner, September 25, 1978. We are grateful to Dr. M. Schimerlik for synthesis of unlabeled PAA. We also thank Drs. J. Knowles and H. Bayley for extensive discussions of the potential uses of azide derivatives as probes of local anesthetic-binding sites. This research was supported by grants from the National Institutes of Health (NS-10294) and from the Muscular Dystrophy Association of America as well as by a National Institutes of Health Predoctoral Fellowship to S.G.B. The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.
Funding AgencyGrant Number
National Institutes of HealthNS-10294
Muscular Dystrophy Association of AmericaUNSPECIFIED
National Institutes of Health Predoctoral FellowshipUNSPECIFIED
Subject Keywords:acetylcholine receptor; photoaffinity labeling; procaine amide azide; histrionicotoxin binding
Issue or Number:1
Record Number:CaltechAUTHORS:BLApnas79
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Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:11495
Deposited By: Tony Diaz
Deposited On:05 Sep 2008 04:59
Last Modified:03 Oct 2019 00:19

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