scholarly journals Interactions of purified bovine brain A1-adenosine receptors with G-proteins. Reciprocal modulation of agonist and antagonist binding

1991 ◽  
Vol 275 (3) ◽  
pp. 651-656 ◽  
Author(s):  
M Freissmuth ◽  
E Selzer ◽  
W Schütz
Keyword(s):  
G Proteins ◽  
Adenosine Receptor ◽  
Adenosine Receptors ◽  
Bovine Brain ◽  
Detergent Solution ◽  
Maximal Effect ◽  
A1 Adenosine Receptor ◽  
Molar Excess ◽  
A1 Adenosine Receptors ◽  
Antagonist Binding

The bovine brain A1-adenosine receptor was purified 8000-fold by affinity chromatography on xanthine-amine-congener (XAC)-Sepharose. Addition of a 120-fold molar excess of a purified bovine brain G-protein preparation (Go,i a mixture of Go and Gi, containing predominantly Go) decreases the Bmax of the binding of the antagonist radioligand [3H]XAC to the receptor. This decrease is observed not only after insertion into phospholipid vesicles but also in detergent solution, and is reversed by GTP analogues. In the presence of Go,i, about 20 and 40% of the receptors display guanine-nucleotide-sensitive high-affinity binding of the agonist radioligand (-)-N6-3-([125I]iodo-4-hydroxyphenylisopropyl)adenosine after reconstitution into lipid vesicles and in detergent solution, respectively. The ability of Go,i to enhance agonist binding and decrease antagonist binding is concentration-dependent, with a half-maximal effect occurring at approximately 10-fold molar excess of G-proteins over A1-adenosine receptors. In the presence of the receptor, the rate of guanosine 5′-[gamma-[35S]thio]triphosphate (GTP[35S]) binding to Go,i is accelerated. This rate is further enhanced if the receptor is activated by the agonist (-)(R)-N6-phenylisopropyladenosine, whereas the antagonist XAC decreases the association rate of GTP[35S] to levels observed in the absence of receptor. These results show (1) that detergent removal is not a prerequisite for the observation of coupling between the A1-adenosine receptor and Go,i, and (2) that the regulatory effect of G-proteins on antagonist binding to the A1-adenosine receptor can be reconstituted by using purified components.

Evidence for regulated coupling of A1 adenosine receptors by phosphorylation in Zucker rats

1995 ◽  
Vol 268 (4) ◽  
pp. E693-E704 ◽  
Author(s):  
D. A. Berkich ◽  
D. R. Luthin ◽  
R. L. Woodard ◽  
S. J. Vannucci ◽  
J. Linden ◽  
...  
Keyword(s):  
Adenosine Receptor ◽  
Adenosine Receptors ◽  
Receptor Agonist ◽  
High Sensitivity ◽  
Zucker Rats ◽  
Guanine Nucleotide ◽  
Agonist Binding ◽  
A1 Adenosine Receptor ◽  
Adenosine Receptor Agonist ◽  
A1 Adenosine Receptors

Studies were designed to find the molecular basis for previous observations that lipolysis is less active and A1 adenosine receptor signaling is more active in adipocytes from obese than from lean Zucker rats. With quantitative immunoblot procedures for detection, Gi alpha 1 and Gs alpha 45 levels were found anomalously low in obese compared with lean membranes (50 and 30%, respectively), but other G alpha subunit levels were normal. However, the sensitivity of the receptor-Gi protein to GTP was about 5- to 10-fold higher in obese compared with lean membranes when assessed from 1) the ability of GTP to inhibit forskolin-stimulated adenylyl cyclase in the presence of an adenosine receptor agonist and 2) the ability of a nonhydrolyzable guanine nucleotide analogue to alter A1 adenosine receptor agonist binding. Alkaline phosphatase treatment of isolated adipocyte membranes from obese but not lean animals decreased guanine nucleotide sensitivity of agonist binding. Surprisingly, solubilized adipocyte A1 adenosine receptors from all animals exhibited the same high sensitivity to guanine nucleotides as that of intact obese membranes, and this high sensitivity could be decreased 20-fold by treatment with alkaline phosphatase. These data suggest that protein phosphorylation may regulate coupling of the A1 adenosine receptor in rat adipocyte membranes.

scholarly journals Focusing on Adenosine Receptors as a Potential Targeted Therapy in Human Diseases

Cells ◽  
2020 ◽  
Vol 9 (3) ◽  
pp. 785 ◽  
Author(s):  
Wiwin Is Effendi ◽  
Tatsuya Nagano ◽  
Kazuyuki Kobayashi ◽  
Yoshihiro Nishimura
Keyword(s):  
G Proteins ◽  
Adenosine Receptor ◽  
Adenosine Receptors ◽  
Signal Pathway ◽  
Human Diseases ◽  
Therapeutic Drug ◽  
Drug Candidates ◽  
Biological Features ◽  
Partial Agonists ◽  
Membrane Bound

Adenosine is involved in a range of physiological and pathological effects through membrane-bound receptors linked to G proteins. There are four subtypes of adenosine receptors, described as A1AR, A2AAR, A2BAR, and A3AR, which are the center of cAMP signal pathway-based drug development. Several types of agonists, partial agonists or antagonists, and allosteric substances have been synthesized from these receptors as new therapeutic drug candidates. Research efforts surrounding A1AR and A2AAR are perhaps the most enticing because of their concentration and affinity; however, as a consequence of distressing conditions, both A2BAR and A3AR levels might accumulate. This review focuses on the biological features of each adenosine receptor as the basis of ligand production and describes clinical studies of adenosine receptor-associated pharmaceuticals in human diseases.

Properties of solubilized and reconstituted A1 adenosine receptors from bovine brain

1991 ◽  
Vol 24 (1) ◽  
pp. 15-23
Author(s):  
Eugenio Ragazzi ◽  
John Shryock ◽  
Krzysztof Palczewski
Keyword(s):  
Adenosine Receptors ◽  
Bovine Brain ◽  
A1 Adenosine Receptors

scholarly journals Heparin and other polyanions uncouple α1-adrenoceptors from G-proteins

1991 ◽  
Vol 280 (3) ◽  
pp. 791-795 ◽  
Author(s):  
L L T Dasso ◽  
C W Taylor
Keyword(s):  
Adenylate Cyclase ◽  
G Protein ◽  
G Proteins ◽  
Alkaline Treatment ◽  
Receptor Activation ◽  
Maximal Effect ◽  
Intact Cells ◽  
High Affinity ◽  
Antagonist Binding ◽  
Coupling Mechanisms

Several polyanionic compounds antagonize the interaction between receptors and the G-proteins that regulate adenylate cyclase or K+ channels, possibly by binding to a basic stretch of the receptor that is proposed to mediate its interaction with the G-proteins. We have studied the effects of polyanions on the interaction between the liver alpha 1-adrenoceptor and the G-protein through which it stimulates polyphosphoinositide turnover. Heparin [concn. causing 50% of maximal effect (EC50) = 0.5 microM], Trypan Blue (EC50 7.1 microM) or suramin (EC50 2.1 microM) prevented formation of the high-affinity adrenaline-receptor-G-protein complex without affecting antagonist binding. After alkaline treatment of the membranes, previously reported to cause G-protein removal, binding of agonists was insensitive to both guanine nucleotides and heparin. We conclude that these polyanions uncouple the alpha 1-adrenoceptor from its G-protein, suggesting that similar coupling mechanisms may underlie receptor activation of the G-proteins that activate polyphosphoinositide hydrolysis and those that regulate adenylate cyclase. This action of heparin severely limits its utility as a selective antagonist of the Ins(1,4,5)P3 receptor in intact cells.

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