摘要：Site-directed mutagenesis was used to investigate the molecular interactions involved in prazosin binding to the human α1b-adrenergic receptor (α1b-AR) receptor. Based on molecular modeling studies, Thr130 and Asp125 in transmembrane region III of the α1b-AR receptor were found to interact with prazosin. Thr130 and Asp125 were mutated to alanine (Ala) and expressed in HEK293 cells. The radioligand [3H]prazosin did not show any binding to Asp125Ala mutant of α1b-AR. Therefore, it was not possible to find any prazosin affinity to the mutant using the radioligand [3H]prazosin. The mutation also abolished phenylephrine-stimulated inositol phosphate (IP) formation of [3H] myo -inositol. On the other hand, the Thr130Ala mutant showed reduced binding affinity for [3H]prazosin (dissociation constant, K d 674.27 p M versus 90.27 p M for the wild-type receptor) and had reduced affinity for both tamsulosin and prazosin (11-fold and 9-fold, respectively). However, the Thr130Ala mutant receptor retained the ability to stimulate the formation of [3H] myo -inositol. The results provide direct evidence that Asp125 and Thr130 are responsible for the interactions between α1b-AR receptor and radioligand [3H]prazosin as well as tamsulosin.
关键词：prazosin;α1b-adrenergic receptor receptor;mutagenesis;binding site
