Functional modifications of the coupling of solubilized dopamine D2 receptors to guanine-nucleotide-binding proteins.
The molecular basis for the regulation of high-affinity agonist, [3H]N-n-propylnorapomorphine ([3H]NPA), binding to cholate-solubilized dopamine D2 receptors was characterized using cations, guanine nucleotides and sulfhydryl-modifying agents. [3H]NPA binding displayed an absolute requirement for divalent cations in the solubilized preparation. Removal of Na+ from the solubilized preparation caused an apparent reconstitution of soluble receptors resulting in a reduced sensitivity of the agonist binding to divalent cations. The pharmacological profile of [3H]NPA binding was found to be similar in membrane and solubilized preparations. N-ethylmaleimide (NEM) and thermal exposure mimicked the effects of guanine nucleotides in reducing the proportion of high-affinity agonist sites in the solubilized state. [3H]NPA binding was much more susceptible to NEM-induced alkylation or heat inactivation compared to the antagonist [3H]spiroperidol binding. Pertussis-toxin-catalyzed ADP-ribosylation of G-proteins in the solubilized preparation resulted in the labelling of only one protein with the apparent molecular weight of 39-41 kDa. Both NEM and heat treatments caused the loss of ADP-ribosylation in the solubilized preparations. A consistent pattern of correlation between receptor binding data and ADP-ribosylation response suggests functional coupling of dopamine D2 receptors to the components of the effector system in solution.
Department of Psychiatry, McMaster University, Hamilton, Ontario, Canada.
S M
SM
Kazmi
L K
LK
Srivastava
R K
RK
Mishra
D000818:Animals; D001058:Apomorphine / Q000031:analogs & derivatives / Q000378:metabolism; D002417:Cattle; D015259:Dopamine Agents / Q000378:metabolism; D005033:Ethylmaleimide / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000032:analysis / Q000378:metabolism* / Q000502:physiology*; D006150:Guanine Nucleotides / Q000494:pharmacology; D006358:Hot Temperature; D008274:Magnesium / Q000494:pharmacology; D008970:Molecular Weight; D037342:Pertussis Toxin; D017448:Receptors, Dopamine D2 / Q000378:metabolism* / Q000502:physiology*; D012964:Sodium / Q000494:pharmacology; D012995:Solubility; D013134:Spiperone / Q000378:metabolism; D014316:Tritium; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D015259:Dopamine Agents; D006150:Guanine Nucleotides; D017448:Receptors, Dopamine D2; D010566:Virulence Factors, Bordetella; D014316:Tritium; D013134:Spiperone; C009074:N-n-propylnorapomorphine; D012964:Sodium; D037342:Pertussis Toxin; D019204:GTP-Binding Proteins; D008274:Magnesium; D001058:Apomorphine; D005033:Ethylmaleimide