Kinetic studies on stereospecific recognition by the thromboxane A2/prostaglandin H2 receptor of the antagonist, S-145.

1. The mechanism for the stereospecific recognition of the antagonist S-145 by the thromboxane A2 (TXA2)/prostaglandin H2 (PGH2) receptor was examined by ligand-binding techniques in rat vascular smooth muscle cells (VSMCs) and in human platelet membranes. 2. Scatchard analysis revealed the existence of a single class of binding sites with the same maximum number for both [3H]-(+)-S-145 and [3H]-(-)-S-145 in both cell types. The dissociation constants (Kd) for the binding of the (+)-isomer in rat VSMCs and human platelet membranes were, respectively, 0.40 +/- 0.03 and 0.20 +/- 0.02 nM, each value being lower than that for the (-)-isomer (3.57 +/- 0.74 and 2.87 +/- 0.08 nM, respectively). 3. The rank orders of potency (Ki) for a series of TXA2/PGH2 ligands at inhibiting [3H]-(+)-S-145 binding were highly correlated with those determined for [3H]-(-)-S-145 binding in both cell preparations. 4. Kinetic analysis of the binding of both radioligands revealed a much lower dissociation rate constant (k-1) and a slightly greater association rate constant (k1) for the (+)-isomer compared to those for the (-)-isomer. 5. These results suggest that it is at the stage of dissociation from the TXA2/PGH2 receptor that the stereochemistry of the optical isomers of S-145 confers their difference in affinity for these receptors in rat VSMCs and human platelet membranes.