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rdf:type | |
lifeskim:mentions | |
pubmed:issue |
2
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pubmed:dateCreated |
1998-12-11
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pubmed:abstractText |
We report the characterization of A2a adenosine receptors (A2aARs) in porcine striatal membranes and their solubilization (25%) by the detergent digitonin. After solubilization, the drug specificity and equilibrium [3H]CGS-21680 ([3H]2-(4-(2-carboxyethyl)phenylethylamino)-5'-N-ethyl-carboxamido -adenosine) binding parameters were virtually identical to those obtained in intact membranes, indicating a conservation of the binding site after the removal of receptors from their lipid environment. Gel filtration on a calibrated Superdex 200 HR column revealed a main [3H]CGS-21680 binding peak with an apparent molecular weight of 171,000+/-9000 Da. In membranes, Scatchard analysis of saturation data carried out in a wide range of radioligand concentration (1-100 nM) resulted in a biphasic curve and, in accordance with the two binding sites model, yielded a Kd1 = 7.4+/-0.5 and Kd2 = 53.1+/-3.6 nM, a Bmax1 = 186+/-15 fmol/mg protein and a Bmax2 = 285+/-20 fmol/mg protein, respectively. In the presence of guanosine-5'-O-(3-thiotriphosphate) (GTPgamma[S]) a shift from two affinity states to a single one was evidenced (Kd = 28.5+/-5.9 nM) and a Bmax value of 504+/-10 fmol/mg protein found. In the soluble extract, only one high-affinity state was detected (Kd = 19.3+/-1.1 nM and Bmax = 285+/-20 fmol/mg protein) and, in the presence of GTPgamma[S]), a two site model likewise provided a significantly (P < 0.01) better fit (Kd1 = 13.9+/-1.2 nM and Kd2 = 72.1+/-6.9 nM, Bmax1 = 125+/-10 fmol/mg protein and Bmax2 = 375+/-19 fmol/mg protein, respectively). These results suggest a close relation between the receptor and G protein solubilized as a functional unit and open the way to its purification.
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pubmed:language |
eng
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pubmed:journal | |
pubmed:citationSubset |
IM
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pubmed:chemical |
http://linkedlifedata.com/resource/pubmed/chemical/2-(4-(2-carboxyethyl)phenethylamino)...,
http://linkedlifedata.com/resource/pubmed/chemical/Adenosine,
http://linkedlifedata.com/resource/pubmed/chemical/Digitonin,
http://linkedlifedata.com/resource/pubmed/chemical/Guanine Nucleotides,
http://linkedlifedata.com/resource/pubmed/chemical/Guanosine 5'-O-(3-Thiotriphosphate),
http://linkedlifedata.com/resource/pubmed/chemical/Phenethylamines,
http://linkedlifedata.com/resource/pubmed/chemical/Receptors, Purinergic P1
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pubmed:status |
MEDLINE
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pubmed:month |
Aug
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pubmed:issn |
0197-0186
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pubmed:author | |
pubmed:issnType |
Print
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pubmed:volume |
33
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pubmed:owner |
NLM
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pubmed:authorsComplete |
Y
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pubmed:pagination |
121-9
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pubmed:dateRevised |
2010-11-18
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pubmed:meshHeading |
pubmed-meshheading:9761456-Adenosine,
pubmed-meshheading:9761456-Animals,
pubmed-meshheading:9761456-Cell Membrane,
pubmed-meshheading:9761456-Chromatography, Gel,
pubmed-meshheading:9761456-Corpus Striatum,
pubmed-meshheading:9761456-Digitonin,
pubmed-meshheading:9761456-Guanine Nucleotides,
pubmed-meshheading:9761456-Guanosine 5'-O-(3-Thiotriphosphate),
pubmed-meshheading:9761456-Molecular Weight,
pubmed-meshheading:9761456-Phenethylamines,
pubmed-meshheading:9761456-Receptors, Purinergic P1,
pubmed-meshheading:9761456-Solubility,
pubmed-meshheading:9761456-Swine
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pubmed:year |
1998
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pubmed:articleTitle |
A2a adenosine receptors: guanine nucleotide derivative regulation in porcine striatal membranes and digitonin soluble fraction.
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pubmed:affiliation |
Dipartimento di Psichiatria, Farmacologia e Biotecnologie dell 'Università degli Studi di Pisa, Italy.
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pubmed:publicationType |
Journal Article,
Research Support, Non-U.S. Gov't
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