Photoaffinity cross-linking of a radioiodinated probe, 125I-A55453, into alpha1-adrenergic receptors
(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.
(Less)
- author
- 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.
- publishing date
- 1984-12-01
- 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
-
- scopus:0021732877
- pmid:6090880
- 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}}, }