Lapetina E G, Reep B, Chang K J
Proc Natl Acad Sci U S A. 1986 Aug;83(16):5880-3. doi: 10.1073/pnas.83.16.5880.
Permeabilization of human platelets with saponin (15-25 micrograms/ml) allows the determination of the ADP-ribosylation of a 41-kDa protein by pertussis toxin. The ADP-ribosylated protein is present in the particulate fraction. ADP-ribosylation of the 41-kDa protein increases for 20 min; it is not affected by indomethacin, prostacyclin, and 1,2-diacylglycerols but is inhibited by 1 mM Ca2+ and phorbol esters. Treatment of platelets with trypsin, thrombin, or collagen before saponin addition precludes subsequent pertussis toxin-induced ADP-ribosylation of the 41-kDa protein. The effect of trypsin or thrombin is blocked by soybean trypsin inhibitor and leupeptin. Trypsin proteolytically cleaves the ADP-ribosylated 41-kDa protein to an ADP-ribosylated fragment slightly smaller than 20 kDa. The results suggest that a modification of a guanine nucleotide-binding regulatory protein is associated with the actions of trypsin, thrombin, and collagen on platelet activation.
用皂角苷（15 - 25微克/毫升）使人类血小板通透化，可测定百日咳毒素对一种41 kDa蛋白质的ADP核糖基化作用。ADP核糖基化的蛋白质存在于颗粒部分。41 kDa蛋白质的ADP核糖基化在20分钟内增加；它不受吲哚美辛、前列环素和1,2 - 二酰甘油的影响，但受到1 mM Ca2+和佛波酯的抑制。在加入皂角苷之前用胰蛋白酶、凝血酶或胶原处理血小板，可阻止随后百日咳毒素诱导的41 kDa蛋白质的ADP核糖基化。胰蛋白酶或凝血酶的作用可被大豆胰蛋白酶抑制剂和亮抑蛋白酶肽阻断。胰蛋白酶将ADP核糖基化的41 kDa蛋白质蛋白水解切割成一个略小于20 kDa的ADP核糖基化片段。结果表明，鸟嘌呤核苷酸结合调节蛋白的一种修饰与胰蛋白酶、凝血酶和胶原对血小板活化的作用有关。
