Solubilization of human platelet alpha-adrenergic receptors: evidence that agonist occupancy of the receptor stabilizes receptor--effector interactions.

Smith, S. Carrie; Limbird, Lee E.

doi:10.1073/pnas.78.7.4026

Cited by 107 publications

(22 citation statements)

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“…p (Lumbird and Lefkowitz, 1978) and w2 (Smith andLumbird, 1981) adrenergic andD, dopaminergic (Kilpatrick andCaron, 1983) wherein high-alfinity agonist binding (which corresponds to the receptor coupled to G protein) is lost subsequent to solubilization. Very scattcred values are currently found in the literature for Kd values of high-and low-affinity states of adenosine receptors in crude membranes (Lohse et al, 1984;Frame et al, 1986;Szot et al, 1987;Rankumar et al, 1988).…”

Section: In Trodu C T I 0 Nmentioning

confidence: 97%

Solubilization of A₁ adenosine receptor from pig brain: Characterization and evidence of the role of the cell membrane on the coexistence of high‐ and low‐affinity states

Casadó

Cantı́

Mallol

et al. 1990

J of Neuroscience Research

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The present solubilization strategy recognizes the important role of detergent cocktails in the solubilization and subsequent stability of adenosine A1, receptors from pig brain cortical membranes. The 3-[3-(cholamidopropyl)dimethylammonio]-1-propane-sulfonate-digitonin mixture produced the extraction of up to 52% of the receptor with an enrichment of 1.2-fold with respect to crude membranes. The binding activity of the soluble extract was very stable even in the absence of glycerol. In crude membranes the existence of high- and low-affinity states was detected, but in the soluble extract and in the detergent-treated membranes only the high-affinity state was detected. Association-dissociation curves showed that in crude membranes no interconversion between high- and low-affinity sites is produced by the association of the ligand [3H]R-N6-phenylisopropyladenosine. These results suggest that the high- and low-affinity states are different conformations induced by the structure of the membrane. The modulation of the binding activity by (Gpp(NH)p) 5'-guanylylimidodiphosphate and Mg2+ was studied. In crude membranes Gpp(NH)p shifted the high-affinity state to the low-affinity state, whereas the contrary occurred when Mg2+ was used. The effect of both Mg2+ and Gpp(NH)p was also assayed with the soluble extract and with the detergent-treated membranes. In addition to a decrease of the overall binding capacity, Gpp(NH)p promoted a conversion to all low-affinity states in the detergent-treated membranes or to all very-low-affinity sites in the soluble extract. Mg2+ and Gpp(NH)p counteracted their effects in intact membranes, whereas Mg2+ could not reverse the uncoupling effect of Gpp(NH)p with solubilized or detergent-treated membranes. Thus, it is suggested that Mg2+ acts at sites other than guanine-nucleotide-sensitive sites. If high-affinity states correspond to receptor/G protein complexes and low-affinity states correspond to the uncoupled receptor, we should conclude that Mg2+, as well as the loss of membrane integrity, favours the interaction of A1 receptor molecule with G protein.

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Section: In Trodu C T I 0 Nmentioning

confidence: 97%

Solubilization of A₁ adenosine receptor from pig brain: Characterization and evidence of the role of the cell membrane on the coexistence of high‐ and low‐affinity states

Casadó

Cantı́

Mallol

et al. 1990

J of Neuroscience Research

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“…In contrast, when P-adrenergic (14), aadrenergic (15), and histamine H1 (16) receptors are solubilized in the absence of agonist, GTP regulation of ligand binding is lost, although ionic effects may be retained for ax-adrenergic (15) and histamine H1 (16) receptors. The GTP binding protein interacts strongly with divalent cations and consists of several subunits (17).…”

mentioning

confidence: 99%

Solubilized Adenosine Receptors in the Brain: Regulation of Guanine Nucleotides

Gavish

Goodman

Snyder

1982

Science

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Adenosine receptors associated with a reduction of adenylate cyclase and labeled by tritium-labeled cyclohexyladenosine can be solubilized from brain membranes with sodium cholate. Regulation of receptor binding by guanine nucleotides is retained in the soluble state. Influences of cations observed in membrane preparations of adenosine receptors are no longer detected with the solubilized receptors. The apparent retention of a complex of receptors and guanosine triphosphate binding but not cation binding protein in the soluble state may permit a molecular analysis of receptor regulation.

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“…Direct evidence of such a process has been provided for the 3-adrenoceptor (Limbird et al, 1 Correspondence. 1980) and the cX2-adrenoceptor (Smith & Limbird, 1981).…”

Section: Introductionmentioning

confidence: 99%

Solubilization and characterization of guanine nucleotide‐sensitive muscarinic agonist binding sites from rat myocardium

Berrie¹,

Birdsall²,

Hulme³

et al. 1984

British J Pharmacology

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Solubilization of human platelet alpha-adrenergic receptors: evidence that agonist occupancy of the receptor stabilizes receptor--effector interactions.

Abstract: The a-adrenergic receptors ofhuman platelet membranes can be directly identified by both a radiolabeled agonist, [3H] Fresh platelet-rich plasma was adjusted to pH 6.5 with citric acid/citrate/dextrose (4). Washed platelet lysates were prepared as described (5)

Cited by 107 publications

References 19 publications

Solubilization of A₁ adenosine receptor from pig brain: Characterization and evidence of the role of the cell membrane on the coexistence of high‐ and low‐affinity states

Solubilization of A₁ adenosine receptor from pig brain: Characterization and evidence of the role of the cell membrane on the coexistence of high‐ and low‐affinity states

Solubilized Adenosine Receptors in the Brain: Regulation of Guanine Nucleotides

Solubilization and characterization of guanine nucleotide‐sensitive muscarinic agonist binding sites from rat myocardium

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Solubilization of human platelet alpha-adrenergic receptors: evidence that agonist occupancy of the receptor stabilizes receptor--effector interactions.

Abstract: The a-adrenergic receptors ofhuman platelet membranes can be directly identified by both a radiolabeled agonist, [3H] Fresh platelet-rich plasma was adjusted to pH 6.5 with citric acid/citrate/dextrose (4). Washed platelet lysates were prepared as described (5)

Cited by 107 publications

References 19 publications

Solubilization of A1 adenosine receptor from pig brain: Characterization and evidence of the role of the cell membrane on the coexistence of high‐ and low‐affinity states

Solubilization of A1 adenosine receptor from pig brain: Characterization and evidence of the role of the cell membrane on the coexistence of high‐ and low‐affinity states

Solubilized Adenosine Receptors in the Brain: Regulation of Guanine Nucleotides

Solubilization and characterization of guanine nucleotide‐sensitive muscarinic agonist binding sites from rat myocardium

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Solubilization of A₁ adenosine receptor from pig brain: Characterization and evidence of the role of the cell membrane on the coexistence of high‐ and low‐affinity states

Solubilization of A₁ adenosine receptor from pig brain: Characterization and evidence of the role of the cell membrane on the coexistence of high‐ and low‐affinity states