Cardin A D, Barnhart R L, Witt K R, Jackson R L
Thromb Res. 1984 Jun 15;34(6):541-50. doi: 10.1016/0049-3848(84)90258-5.
The heparin-binding properties of human plasma apolipoproteins B-100 and E (apoB-100 and E) of low density lipoproteins (LDL), thrombin, and antithrombin-III (AT-III) were investigated. A highly reactive heparin (HRH) to apoB-100 was isolated by chromatography of crude heparin on a column of LDL immobilized to Affi-Gel 10. This HRH showed a high, Ca2+-dependent precipitating activity towards LDL; 1 microgram HRH uronic acid precipitated 50-70 micrograms LDL-protein. HRH was fractionated further by chromatography on a column of AT-III bound to concanavalin A-Sepharose. The unretained fraction of heparin (HRH1) had a low affinity for AT-III. The bound heparin (HRH2) had a high affinity for AT-III and precipitated LDL in the presence of Ca2+. To assess further their heparin-binding properties, the proteins were subjected to gradient-gel electrophoresis under denaturing conditions, transferred to nitrocellulose by electrophoresis, and then assayed for their ability to bind [125I]-labeled HRH2. Autoradiographic analysis showed that thrombin, apolipoproteins E and B-100, and the AT-III . thrombin covalent complex bound HRH2. Denatured AT-III did not bind HRH2, indicating that its heparin recognition site may depend on conformation.
研究了人血浆低密度脂蛋白（LDL）中的载脂蛋白B - 100和E（apoB - 100和E）、凝血酶及抗凝血酶III（AT - III）的肝素结合特性。通过将粗肝素在固定于Affi - Gel 10的LDL柱上进行层析，分离出一种对apoB - 100具有高反应性的肝素（HRH）。这种HRH对LDL表现出高的、Ca2 +依赖的沉淀活性；1微克HRH糖醛酸可沉淀50 - 70微克LDL蛋白。通过在结合伴刀豆球蛋白A - 琼脂糖的AT - III柱上进行层析，对HRH进一步分级分离。未保留的肝素级分（HRH1）对AT - III的亲和力较低。结合的肝素（HRH2）对AT - III具有高亲和力，并在Ca2 +存在下沉淀LDL。为进一步评估它们的肝素结合特性，在变性条件下对这些蛋白质进行梯度凝胶电泳，通过电泳转移至硝酸纤维素膜上，然后检测它们结合[125I]标记的HRH2的能力。放射自显影分析表明，凝血酶、载脂蛋白E和B - 100以及AT - III - 凝血酶共价复合物结合HRH2。变性的AT - III不结合HRH2，表明其肝素识别位点可能取决于构象。
