Katada T, Bokoch G M, Northup J K, Ui M, Gilman A G
J Biol Chem. 1984 Mar 25;259(6):3568-77.
Treatment of membranes with islet activating protein (IAP), a toxin from Bordetella pertussis, results in abolition of GTP-dependent, receptor-mediated inhibition of adenylate cyclase. This appears to result from IAP-catalyzed ADP-ribosylation of a 41,000-Da membrane-bound protein. A protein with 41,000- and 35,000-Da subunits has been purified from rabbit liver membranes as the predominant substrate for IAP. This protein has now been shown to be capable of regulating membrane-bound adenylate cyclase activity of human platelets under various conditions. The characteristics of the actions of the IAP substrate are as follows. 1) Purified 41,000/35,000-Da dimer is capable of restoring the inhibitory effects of guanine nucleotides and the alpha 2-adrenergic agonist, epinephrine, on the adenylate cyclase activity of IAP-treated membranes. 2) The subunits of the dimer dissociate in the presence of guanine nucleotide analogs or A1(3+), Mg2+, and F-. The 41,000-Da subunit has a high affinity binding site for guanine nucleotides. 3) The resolved 35,000-Da subunit of the dimer mimics guanine nucleotide- and epinephrine-induced inhibition of adenylate cyclase. 4) The resolved (unliganded) 41,000-Da subunit stimulates adenylate cyclase activity and relieves guanine nucleotide- +/- epinephrine-induced inhibition of the enzyme. In contrast, the GTP gamma S-bound form of the 41,000-Da subunit inhibits adenylate cyclase activity, although with lower apparent affinity than does the 35,000-Da subunit. 5) The 35,000-Da subunit increases the rate of deactivation of Gs, the stimulatory regulatory protein of adenylate cyclase. In contrast, the 41,000-Da subunit can interact with Gs and inhibit its deactivation. These data strongly suggest that the IAP substrate is another dimeric, guanine nucleotide-binding regulatory protein and that it is responsible for inhibitory modulation of adenylate cyclase activity.
用百日咳博德特氏菌毒素胰岛激活蛋白（IAP）处理细胞膜，会导致GTP依赖性、受体介导的腺苷酸环化酶抑制作用消失。这似乎是由于IAP催化一种41,000道尔顿的膜结合蛋白进行ADP核糖基化所致。一种由41,000道尔顿和35,000道尔顿亚基组成的蛋白已从兔肝细胞膜中纯化出来，作为IAP的主要底物。现已证明该蛋白能够在各种条件下调节人血小板的膜结合腺苷酸环化酶活性。IAP底物的作用特性如下：1）纯化的41,000/35,000道尔顿二聚体能够恢复鸟嘌呤核苷酸和α2 -肾上腺素能激动剂肾上腺素对IAP处理细胞膜的腺苷酸环化酶活性的抑制作用。2）二聚体的亚基在鸟嘌呤核苷酸类似物或Al(3+)、Mg2+和F -存在时会解离。41,000道尔顿的亚基具有一个对鸟嘌呤核苷酸有高亲和力的结合位点。3）二聚体中分离出的35,000道尔顿亚基模拟鸟嘌呤核苷酸和肾上腺素诱导的腺苷酸环化酶抑制作用。4）分离出的（未结合配体的）41,000道尔顿亚基刺激腺苷酸环化酶活性，并解除鸟嘌呤核苷酸±肾上腺素诱导的该酶抑制作用。相比之下，41,000道尔顿亚基的GTPγS结合形式抑制腺苷酸环化酶活性，尽管其表观亲和力低于35,000道尔顿亚基。5）35,000道尔顿亚基增加了腺苷酸环化酶刺激调节蛋白Gs的失活速率。相比之下，41,000道尔顿亚基可与Gs相互作用并抑制其失活。这些数据有力地表明，IAP底物是另一种二聚体鸟嘌呤核苷酸结合调节蛋白，并且它负责对腺苷酸环化酶活性的抑制性调节。
