Bu G, Williams S, Strickland D K, Schwartz A L
Edward Mallinckrodt Department of Pediatrics, Washington University School of Medicine, St. Louis, MO 63110.
Proc Natl Acad Sci U S A. 1992 Aug 15;89(16):7427-31. doi: 10.1073/pnas.89.16.7427.
Tissue-type plasminogen activator (t-PA), a serine protease that catalyzes the initial and rate-limiting step in the fibrinolytic cascade, is cleared rapidly in vivo by the liver. Using chemical crosslinking, we have recently identified a plasminogen-activator inhibitor type 1 (PAI-1)-independent t-PA clearance receptor on rat hepatoma MH1C1 cells with a relative molecular mass of approximately 500 kDa. Another recently identified membrane receptor, low density lipoprotein receptor-related protein/alpha 2-macroglobulin receptor (LRP/alpha 2MR), was also detected on MH1C1 hepatoma cells by using immunoprecipitation with anti-LRP/alpha 2MR antibody. When analyzed by SDS/PAGE, we found the t-PA receptor identified on MH1C1 cells comigrated with the large subunit of LRP/alpha 2MR. The t-PA receptor was immunoprecipitated by an anti-LRP/alpha 2MR antibody after chemical crosslinking of specifically bound 125I-labeled t-PA to its receptor. Through chemical crosslinking studies, we found that t-PA and methylamine-activated alpha 2-macroglobulin could bind to LRP/alpha 2MR simultaneously without competing with one another for binding, suggesting that the two ligands bound to two independent sites on the LRP/alpha 2MR molecule. Furthermore, a 39-kDa protein, which modulates ligand binding to LRP/alpha 2MR, was also found to inhibit t-PA binding to its receptor. These data thus show that the t-PA clearance receptor identified on MH1C1 hepatoma cells is LRP/alpha 2MR.
组织型纤溶酶原激活剂(t-PA)是一种丝氨酸蛋白酶,可催化纤维蛋白溶解级联反应的起始和限速步骤,在体内会被肝脏迅速清除。我们最近利用化学交联技术,在大鼠肝癌MH1C1细胞上鉴定出一种相对分子质量约为500 kDa的纤溶酶原激活剂抑制剂1型(PAI-1)非依赖性t-PA清除受体。通过用抗LRP/α2MR抗体进行免疫沉淀,在MH1C1肝癌细胞上还检测到了另一种最近鉴定出的膜受体,即低密度脂蛋白受体相关蛋白/α2巨球蛋白受体(LRP/α2MR)。当通过SDS/PAGE分析时,我们发现MH1C1细胞上鉴定出的t-PA受体与LRP/α2MR的大亚基迁移率相同。在将特异性结合的125I标记的t-PA与其受体进行化学交联后,t-PA受体可被抗LRP/α2MR抗体免疫沉淀。通过化学交联研究,我们发现t-PA和甲胺激活的α2巨球蛋白可以同时结合到LRP/α2MR上,而不会相互竞争结合,这表明这两种配体结合到LRP/α2MR分子上的两个独立位点。此外,还发现一种调节配体与LRP/α2MR结合的39 kDa蛋白也能抑制t-PA与其受体的结合。因此,这些数据表明在MH1C1肝癌细胞上鉴定出的t-PA清除受体是LRP/α2MR。