The kyotorphin (tyrosine-arginine) receptor and a selective reconstitution with purified G(i), measured with GTPase and phospholipase C assays

H. Ueda, Y. Yoshihara, H. Misawa, N. Fukushima, T. Katada, M. Ui, H. Takagi, M. Satoh

研究成果: Article査読

66 被引用数 (Scopus)

抄録

We attempted to identify the kyotorphin receptor and the post receptor mechanisms mediated by GTP-binding proteins (G-proteins), using reconstitution techniques. The specific binding of [3H]kyotorphin in rat brain membranes was composed of high affinity (K(d) = 0.34 nM) and low affinity (K(d) = 9.07 nM) binding. As the high affinity binding disappeared in the presence of guanosine 5'-O-(3-thiotriphosphate) and MgCl2, we investigated the kyotorphin receptor-mediated changes in membrane G-protein activity by measuring low K(m) GTPase activity. Kyotorphin produced a stimulation of low K(m) GTPase, and this stimulation was antagonized by Leu-Arg, a synthetic dipeptide which showed a potent displacement of [3H]kyotorphin binding, yet in itself had no effect on the low K(m) GTPase. The kyotorphin stimulation of low K(m) GTPase was abolished by pretreating membranes with islet-activating protein, pertussis toxin, and was recovered by reconstitution with purified G-protein, G(i), but not with G(o). Similar evidence of selective coupling of kyotorphin receptor to G(i) was obtained with the phospholipase C assay. Kyotorphin-induced stimulation of phospholipase C was also abolished by islet-activating protein-treatment and recovered by reconstitution with G(i) but not with G(o). These findings indicate that specific high and low affinity kyotorphin receptors exist in the rat brain and that the kyotorphin receptor is functionally coupled to stimulation of phospholipase C, through G(i). This study provides the first evidence of a selective involvement of G(i) in the receptor-mediated activation of phospholipase C.

本文言語English
ページ(範囲)3732-3741
ページ数10
ジャーナルJournal of Biological Chemistry
264
7
出版ステータスPublished - 1989
外部発表はい

ASJC Scopus subject areas

  • 生化学
  • 分子生物学
  • 細胞生物学

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