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Photoaffinity labeling of the DDT1 MF-2 cell α1-adrenergic receptor

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Summary

In this study, we have used an α1-adrenergic receptor photoaffinity ligand, 2-[4-(4-azido-3-iodo-benzoyl)-piperazin-1-yl]-4-amino-6, 7-dimethoxyquinazoline (125I-APD), to label covalently the α1-adrenergic receptor in a smooth muscle cell line. Our results indicate that in the absence of light, (125I)APD binds reversibly to a site in the DDT1 MF-2 cell membranes having pharmacological characteristics of an α1-adrenergic receptor. Following incorporation of (125I)ADP into partially purified membranes a single labeled band of protein with a Mr of 81 000 was visualized by autoradiography following sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Incorporation of (125I)-APD into this band was affected by adrenergic agonists and antagonists in a manner consistent with an α1-adrenergic interaction. Prazosin (α1-selective) blocked incorporation of the label into the Mr = 81 000 protein while yohimbine (α2-selective) did not. Of the adrenergic agonists, (−)-epinephrine and (−)-norepinephrine but not (−)-isoproterenol blocked labeling of the Mr − 81 000 protein. We conclude that the ligand binding site of the DDT1 MF-2 cell α1-adrenergic receptor resides in a Mr = 81 000 protein.

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Authors and Affiliations

  1. Departments of Physiology and Biophysics, University of Arkansas for Medical Sciences, 4301 West Markham Street, 72205, Little Rock, Arkansas, USA

    Lawrence E. Cornett Ph.D.

  2. Departments of Medicine, University of Arkansas for Medical Sciences, 4301 West Markham Street, 72205, Little Rock, Arkansas, USA

    James S. Norris

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  1. Lawrence E. Cornett Ph.D.
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  2. James S. Norris
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Cornett, L.E., Norris, J.S. Photoaffinity labeling of the DDT1 MF-2 cell α1-adrenergic receptor. Mol Cell Biochem 67, 47–53 (1985). https://doi.org/10.1007/BF00220985

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  • DOI: https://doi.org/10.1007/BF00220985

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