Thermodynamics of full agonist, partial agonist, and antagonist binding to wild-type and mutant adenosine A1 receptors

Biochem Pharmacol. 1998 Dec 1;56(11):1437-45. doi: 10.1016/s0006-2952(98)00202-0.


A thermodynamic analysis of the binding of a full agonist (N6-cyclopentyladenosine), a partial agonist (8-butylamino-N6-cyclopentyladenosine) and an antagonist (8-cyclopentyltheophylline) to human wild-type and mutant (mutation of a threonine (Thr) to an alanine (Ala) residue at position 277) adenosine A1 receptors expressed on Chinese hamster ovary (CHO) cells, and to rat brain adenosine A1 receptors was undertaken. The thermodynamic parameters deltaGo (standard free energy), deltaHo (standard enthalpy) and deltaSo (standard entropy) of the binding equilibrium to rat brain receptors were determined by means of affinity measurements carried out at four different temperatures (0, 10, 20 and 25 degrees) and van't Hoff plots. Two temperatures (0 and 25 degrees) were considered for human receptors. Affinity constants were obtained from inhibition assays on membrane preparations of rat brain and CHO cells by use of the antagonist [3H]1,3-dipropyl-8-cyclopentylxanthine ([3H]DPCPX) as selective adenosine A1 receptor radioligand. As for rat brain receptors, full agonist binding was totally entropy driven, whereas antagonist binding was essentially enthalpy driven. Partial agonist binding appeared both enthalpy and entropy driven. As for human receptors, full agonist affinity was highly dependent on the presence of Thr277. Moreover, affinity to both wild-type and mutant receptors was enhanced by temperature increase, suggesting a totally entropy-driven binding. Antagonist binding did not depend on the presence of Thr277. Antagonist affinity decreased with an increase in temperature, suggesting a mainly enthalpy-driven binding. Partial agonist binding was significantly dependent on the presence of Thr277 at 25 degrees, whereas such a dependence was not evident at 0 degrees. It is concluded that Thr277 contributes only to the binding of adenosine derivatives and that its role changes drastically with the receptor conformation and with the type of agonist (full or partial) interacting with the adenosine A1 receptors.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine / analogs & derivatives*
  • Adenosine / chemistry
  • Adenosine / metabolism
  • Alanine
  • Amino Acid Substitution
  • Animals
  • Binding, Competitive
  • CHO Cells
  • Cricetinae
  • Entropy
  • Humans
  • Kinetics
  • Point Mutation
  • Purinergic P1 Receptor Agonists
  • Purinergic P1 Receptor Antagonists
  • Radioligand Assay
  • Rats
  • Receptors, Purinergic P1 / chemistry*
  • Receptors, Purinergic P1 / metabolism*
  • Theophylline / analogs & derivatives*
  • Theophylline / chemistry
  • Theophylline / metabolism
  • Thermodynamics
  • Threonine
  • Transfection
  • Xanthines / metabolism


  • 8-butylamino-N(6)-cyclopentyladenosine
  • Purinergic P1 Receptor Agonists
  • Purinergic P1 Receptor Antagonists
  • Receptors, Purinergic P1
  • Xanthines
  • Threonine
  • 8-cyclopentyl-1,3-dimethylxanthine
  • N(6)-cyclopentyladenosine
  • 1,3-dipropyl-8-cyclopentylxanthine
  • Theophylline
  • Adenosine
  • Alanine