CORRELATION BETWEEN GUANINE-NUCLEOTIDE EFFECT AND REVERSIBLE BINDING PROPERTY OF ENDOTHELIN ANALOGS

[I-125]-IRL-1620 and [I-125]-ET-1 (readily reversible and essentially irreversible endothelin (ET) receptor agonists, respectively) were use d to demonstrate the relationship between the reversible binding natur e of ET receptor agonists and guanine nucleotide effect using ET(B) re ceptors as the model system. Addition of increasing concentrations of GTP(gamma)s to membranes prepared from Chinese hamster ovary (CHO) cel ls stably transfected with human ET(B) receptors, dog lung and pig lun g decreased [I-125]-IRL-1620 binding to these membranes between 50% an d 60%, whereas [I-125]-ET-1 binding to these receptors was unaffected by GTP(gamma)s. Saturation binding experiments in the absence and pres ence of 100 mu M GTP(gamma)s indicated that the apparent dissociation constant [K-d(apparent)] for [I-125]-IRL-1620 was increased 2 to 2.4-f old in all 3 membrane preparations in the presence of GTP(gamma)s comp ared to its absence. There was no difference in the apparent dissociat ion constants of [I-125]-ET-1 in the presence and absence of GTP gamma s in these membrane preparations. This inhibitory effect was specific for guanosine triphosphate since adenine nucleotides failed to decrea se the affinity of [I-125]-IRL-1620 for the receptors. The correlation between guanine nucleotide effect and reversible binding property of the agonist was further strengthened by the observation that in rat ce rebellum and rat renal papilla, where [I-125]-IRL-1620 binding was irr eversible, guanine nucleotides had no effect on the binding of this li gand. These data clearly indicate that there is a good correlation bet ween the reversible binding property of the ET receptor agonist and th e guanine nucleotide effect on the binding of the agonist.