Dixit V M, Grant G A, Frazier W A, Santoro S A
Biochem Biophys Res Commun. 1984 Mar 30;119(3):1075-81. doi: 10.1016/0006-291x(84)90884-2.
Purified platelet thrombospondin binds to immobilized fibrinogen if both Ca++ and Mg++ are present. Digestion of the purified molecule with thermolysin results in a limited number of discrete proteolytic fragments. When such digests are subjected to affinity chromatography on immobilized fibrinogen, only the fragments with Mr of 120,000 and 140,000 are specifically bound and subsequently eluted by the addition of EDTA to the column buffer. Examination by SDS-PAGE under both reducing and nonreducing conditions reveals that the fibrinogen-binding domain is derived from the region of the thrombospondin molecule containing the interchain disulfide bonds. The requirement for Ca++ and Mg++ for optimal binding to fibrinogen is also manifest by the Mr 120,000/140,000 thermolytic fragments.
如果同时存在钙离子（Ca++）和镁离子（Mg++），纯化的血小板凝血酶敏感蛋白会与固定化的纤维蛋白原结合。用嗜热菌蛋白酶消化纯化的分子会产生数量有限的离散蛋白水解片段。当将这些消化产物在固定化纤维蛋白原上进行亲和层析时，只有分子量为120,000和140,000的片段会特异性结合，随后通过向柱缓冲液中添加乙二胺四乙酸（EDTA）将其洗脱。在还原和非还原条件下通过十二烷基硫酸钠 - 聚丙烯酰胺凝胶电泳（SDS - PAGE）检测发现，纤维蛋白原结合结构域源自凝血酶敏感蛋白分子中包含链间二硫键的区域。分子量为120,000/140,000的嗜热菌蛋白酶消化片段也表现出对钙离子（Ca++）和镁离子（Mg++）与纤维蛋白原最佳结合的需求。
