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Photoaffinity cross-linking of a radioiodinated probe, 125I-A55453, into alpha1-adrenergic receptors

Dickinson, K. E.J. ; Leeb-Lundberg, L. M.F. LU ; Heald, S. L. ; Wikberg, J. E. ; DeBernardis, J. F. ; Caron, M. G. and Lefkowitz, R. J. (1984) In Molecular Pharmacology 26(2). p.187-195
Abstract

We have synthesized and characterized a high-affinity alpha1-adrenergic receptor probe, 4-amino-6,7-dimethoxy-2[4'-[5''-(3-125I-iodo-4-aminophenyl)pentan oyl]-1'-piperazinyl]quinazoline (125I-A55453). This ligand binds reversibly to rat hepatic plasma membranes with high affinity (K(D) = 77 ± 6 pM), and it labels the same number of 'specific' prazosin-competable sites as the alpha1-adrenergic receptor-selective radioligand [125I] iodo-2-[β-(4-hydroxyphenyl)-ethylaminomethyl]tetralone. Specific binding is stereoselective and competed for by alpha-adrenergic agents with an alpha1-adrenergic receptor specificity. 125I-A55453 can be covalently photoincorporated... (More)

We have synthesized and characterized a high-affinity alpha1-adrenergic receptor probe, 4-amino-6,7-dimethoxy-2[4'-[5''-(3-125I-iodo-4-aminophenyl)pentan oyl]-1'-piperazinyl]quinazoline (125I-A55453). This ligand binds reversibly to rat hepatic plasma membranes with high affinity (K(D) = 77 ± 6 pM), and it labels the same number of 'specific' prazosin-competable sites as the alpha1-adrenergic receptor-selective radioligand [125I] iodo-2-[β-(4-hydroxyphenyl)-ethylaminomethyl]tetralone. Specific binding is stereoselective and competed for by alpha-adrenergic agents with an alpha1-adrenergic receptor specificity. 125I-A55453 can be covalently photoincorporated into peptides of rat hepatic and splenic membranes using the bifunctional photoactive cross-linker, N-succinimidyl-6-(4'-azido-2'-nitrophenylamino)hexanoate. Following photolysis, sodium dodecyl sulfate-polyacrylamide gel electrophoresis of labeled hepatic membranes reveals a major 'specifically' labeled peptide of M(r) = 82,000 (± 1,000) with minor peptides at M(r) = 50,000 (± 500), and 40,000 (± 300). Covalent incorporation of 125I-A55453 into the M(r) = 82,000 peptide is inhibited by adrenergic drugs with an alpha1-adrenergic receptor specificity. Labeled splenic membranes demonstrate a broad band of photoincorporated radioactivity centered at M(r) = 82,000, and covalent incorporation into this peptide is also attenuated with an alpha1-adrenergic receptor specificity. This new high-affinity radioiodinated probe has features which should make it useful for the molecular characterization of alpha1-adrenergic receptors in tissues.

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author
; ; ; ; ; and
publishing date
type
Contribution to journal
publication status
published
subject
in
Molecular Pharmacology
volume
26
issue
2
pages
187 - 195
publisher
American Society for Pharmacology and Experimental Therapeutics
external identifiers
  • pmid:6090880
  • scopus:0021732877
ISSN
0026-895X
language
English
LU publication?
no
id
b29ced02-63ed-4b95-ae95-bd60588a679e
date added to LUP
2019-06-04 14:47:17
date last changed
2024-01-01 09:18:36
@article{b29ced02-63ed-4b95-ae95-bd60588a679e,
  abstract     = {{<p>We have synthesized and characterized a high-affinity alpha<sub>1</sub>-adrenergic receptor probe, 4-amino-6,7-dimethoxy-2[4'-[5''-(3-<sup>125</sup>I-iodo-4-aminophenyl)pentan oyl]-1'-piperazinyl]quinazoline (<sup>125</sup>I-A55453). This ligand binds reversibly to rat hepatic plasma membranes with high affinity (K(D) = 77 ± 6 pM), and it labels the same number of 'specific' prazosin-competable sites as the alpha<sub>1</sub>-adrenergic receptor-selective radioligand [<sup>125</sup>I] iodo-2-[β-(4-hydroxyphenyl)-ethylaminomethyl]tetralone. Specific binding is stereoselective and competed for by alpha-adrenergic agents with an alpha<sub>1</sub>-adrenergic receptor specificity. <sup>125</sup>I-A55453 can be covalently photoincorporated into peptides of rat hepatic and splenic membranes using the bifunctional photoactive cross-linker, N-succinimidyl-6-(4'-azido-2'-nitrophenylamino)hexanoate. Following photolysis, sodium dodecyl sulfate-polyacrylamide gel electrophoresis of labeled hepatic membranes reveals a major 'specifically' labeled peptide of M(r) = 82,000 (± 1,000) with minor peptides at M(r) = 50,000 (± 500), and 40,000 (± 300). Covalent incorporation of <sup>125</sup>I-A55453 into the M(r) = 82,000 peptide is inhibited by adrenergic drugs with an alpha<sub>1</sub>-adrenergic receptor specificity. Labeled splenic membranes demonstrate a broad band of photoincorporated radioactivity centered at M(r) = 82,000, and covalent incorporation into this peptide is also attenuated with an alpha<sub>1</sub>-adrenergic receptor specificity. This new high-affinity radioiodinated probe has features which should make it useful for the molecular characterization of alpha<sub>1</sub>-adrenergic receptors in tissues.</p>}},
  author       = {{Dickinson, K. E.J. and Leeb-Lundberg, L. M.F. and Heald, S. L. and Wikberg, J. E. and DeBernardis, J. F. and Caron, M. G. and Lefkowitz, R. J.}},
  issn         = {{0026-895X}},
  language     = {{eng}},
  month        = {{12}},
  number       = {{2}},
  pages        = {{187--195}},
  publisher    = {{American Society for Pharmacology and Experimental Therapeutics}},
  series       = {{Molecular Pharmacology}},
  title        = {{Photoaffinity cross-linking of a radioiodinated probe, <sup>125</sup>I-A55453, into alpha<sub>1</sub>-adrenergic receptors}},
  volume       = {{26}},
  year         = {{1984}},
}