K. Klotz, G. Cristalli, M. Grifantini, S. Vittori, und M. Lohse. J Biol Chem, 260 (27):
14659-64(November 1985)Klotz, K N Cristalli, G Grifantini, M Vittori, S Lohse, M J Research
Support, Non-U.S. Gov't United states The Journal of biological chemistry
J Biol Chem. 1985 Nov 25;260(27):14659-64..
Zusammenfassung
The ligand-binding subunit of the A1-adenosine receptor has been identified
by photoaffinity labeling. A photolabile derivative of R-N6-phenylisopropyladenosine,
R-2-azido-N6-p-hydroxyphenylisopropyladenosine (R-AHPIA), has been
synthesized as a covalent specific ligand for A1-adenosine receptors.
In adenylate cyclase studies with membranes of rat fat cells and
human platelets, R-AHPIA has adenosine receptor agonist activity
with a more than 60-fold selectivity for the A1-subtype. It competes
for 3HN6-phenylisopropyladenosine binding to A1-receptors of rat
brain membranes with a Ki value of 1.6 nM. After UV irradiation,
R-AHPIA binds irreversibly to the receptor, as indicated by a loss
of 3HN6-phenylisopropyladenosine binding after extensive washing;
the Ki value for this photoinactivation is 1.3 nM. The p-hydroxyphenyl
substituent of R-AHPIA can be directly radioiodinated to give a photoaffinity
label of high specific radioactivity (125I-AHPIA). This compound
has a KD value of about 1.5 nM as assessed from saturation and kinetic
experiments. Adenosine analogues compete for 125I-AHPIA binding to
rat brain membranes with an order of potency characteristic for A1-adenosine
receptors. Dissociation curves following UV irradiation at equilibrium
demonstrate 30-40% irreversible specific binding. Sodium dodecyl
sulfate-polyacrylamide gel electrophoresis indicates that the probe
is photoincorporated into a single peptide of Mr = 35,000. Labeling
of this peptide can be blocked specifically and stereoselectively
by adenosine receptor agonists and antagonists in a manner which
is typical for the A1-subtype. The results indicate that 125I-AHPIA
identifies the ligand-binding subunit of the A1-adenosine receptor,
which is a peptide with Mr = 35,000.
Klotz, K N Cristalli, G Grifantini, M Vittori, S Lohse, M J Research
Support, Non-U.S. Gov't United states The Journal of biological chemistry
J Biol Chem. 1985 Nov 25;260(27):14659-64.
%0 Journal Article
%1 Klotz1985
%A Klotz, K. N.
%A Cristalli, G.
%A Grifantini, M.
%A Vittori, S.
%A Lohse, M. J.
%D 1985
%J J Biol Chem
%K & Adenosine/*analogs Adenylate Adipose Affinity Animals Azides/*pharmacology Blood Brain/metabolism Cell Cyclase/metabolism Humans Iodine Kinetics Labels/*pharmacology Membrane/metabolism Phenylisopropyladenosine/*analogs Platelets/metabolism Purinergic Radioisotopes Rats Surface/drug Tissue/metabolism derivatives/*metabolism derivatives/pharmacology effects/*metabolism Receptor
%N 27
%P 14659-64
%T Photoaffinity labeling of A1-adenosine receptors
%U http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=2997218
%V 260
%X The ligand-binding subunit of the A1-adenosine receptor has been identified
by photoaffinity labeling. A photolabile derivative of R-N6-phenylisopropyladenosine,
R-2-azido-N6-p-hydroxyphenylisopropyladenosine (R-AHPIA), has been
synthesized as a covalent specific ligand for A1-adenosine receptors.
In adenylate cyclase studies with membranes of rat fat cells and
human platelets, R-AHPIA has adenosine receptor agonist activity
with a more than 60-fold selectivity for the A1-subtype. It competes
for 3HN6-phenylisopropyladenosine binding to A1-receptors of rat
brain membranes with a Ki value of 1.6 nM. After UV irradiation,
R-AHPIA binds irreversibly to the receptor, as indicated by a loss
of 3HN6-phenylisopropyladenosine binding after extensive washing;
the Ki value for this photoinactivation is 1.3 nM. The p-hydroxyphenyl
substituent of R-AHPIA can be directly radioiodinated to give a photoaffinity
label of high specific radioactivity (125I-AHPIA). This compound
has a KD value of about 1.5 nM as assessed from saturation and kinetic
experiments. Adenosine analogues compete for 125I-AHPIA binding to
rat brain membranes with an order of potency characteristic for A1-adenosine
receptors. Dissociation curves following UV irradiation at equilibrium
demonstrate 30-40% irreversible specific binding. Sodium dodecyl
sulfate-polyacrylamide gel electrophoresis indicates that the probe
is photoincorporated into a single peptide of Mr = 35,000. Labeling
of this peptide can be blocked specifically and stereoselectively
by adenosine receptor agonists and antagonists in a manner which
is typical for the A1-subtype. The results indicate that 125I-AHPIA
identifies the ligand-binding subunit of the A1-adenosine receptor,
which is a peptide with Mr = 35,000.
@article{Klotz1985,
abstract = {The ligand-binding subunit of the A1-adenosine receptor has been identified
by photoaffinity labeling. A photolabile derivative of R-N6-phenylisopropyladenosine,
R-2-azido-N6-p-hydroxyphenylisopropyladenosine (R-AHPIA), has been
synthesized as a covalent specific ligand for A1-adenosine receptors.
In adenylate cyclase studies with membranes of rat fat cells and
human platelets, R-AHPIA has adenosine receptor agonist activity
with a more than 60-fold selectivity for the A1-subtype. It competes
for [3H]N6-phenylisopropyladenosine binding to A1-receptors of rat
brain membranes with a Ki value of 1.6 nM. After UV irradiation,
R-AHPIA binds irreversibly to the receptor, as indicated by a loss
of [3H]N6-phenylisopropyladenosine binding after extensive washing;
the Ki value for this photoinactivation is 1.3 nM. The p-hydroxyphenyl
substituent of R-AHPIA can be directly radioiodinated to give a photoaffinity
label of high specific radioactivity (125I-AHPIA). This compound
has a KD value of about 1.5 nM as assessed from saturation and kinetic
experiments. Adenosine analogues compete for 125I-AHPIA binding to
rat brain membranes with an order of potency characteristic for A1-adenosine
receptors. Dissociation curves following UV irradiation at equilibrium
demonstrate 30-40% irreversible specific binding. Sodium dodecyl
sulfate-polyacrylamide gel electrophoresis indicates that the probe
is photoincorporated into a single peptide of Mr = 35,000. Labeling
of this peptide can be blocked specifically and stereoselectively
by adenosine receptor agonists and antagonists in a manner which
is typical for the A1-subtype. The results indicate that 125I-AHPIA
identifies the ligand-binding subunit of the A1-adenosine receptor,
which is a peptide with Mr = 35,000.},
added-at = {2010-12-14T18:12:02.000+0100},
author = {Klotz, K. N. and Cristalli, G. and Grifantini, M. and Vittori, S. and Lohse, M. J.},
biburl = {https://www.bibsonomy.org/bibtex/21959d970d5f301aa6b6d5185eb9ce4eb/pharmawuerz},
endnotereftype = {Journal Article},
interhash = {bc2483a0b6baddb1689373e28a97765f},
intrahash = {1959d970d5f301aa6b6d5185eb9ce4eb},
issn = {0021-9258 (Print) 0021-9258 (Linking)},
journal = {J Biol Chem},
keywords = {& Adenosine/*analogs Adenylate Adipose Affinity Animals Azides/*pharmacology Blood Brain/metabolism Cell Cyclase/metabolism Humans Iodine Kinetics Labels/*pharmacology Membrane/metabolism Phenylisopropyladenosine/*analogs Platelets/metabolism Purinergic Radioisotopes Rats Surface/drug Tissue/metabolism derivatives/*metabolism derivatives/pharmacology effects/*metabolism Receptor},
month = {Nov 25},
note = {Klotz, K N Cristalli, G Grifantini, M Vittori, S Lohse, M J Research
Support, Non-U.S. Gov't United states The Journal of biological chemistry
J Biol Chem. 1985 Nov 25;260(27):14659-64.},
number = 27,
pages = {14659-64},
shorttitle = {Photoaffinity labeling of A1-adenosine receptors},
timestamp = {2010-12-14T18:20:04.000+0100},
title = {Photoaffinity labeling of A1-adenosine receptors},
url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=2997218},
volume = 260,
year = 1985
}