Calzada Maria J, Kuznetsova Svetlana A, Sipes John M, Rodrigues Rui G, Cashel Jo Anne, Annis Douglas S, Mosher Deane F, Roberts David D
Laboratory of Pathology, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, United States.
Matrix Biol. 2008 May;27(4):339-51. doi: 10.1016/j.matbio.2007.12.002. Epub 2007 Dec 15.
Conformational changes induced in thrombospondin-1 by removal of calcium regulate interactions with some ligands of its N-modules. Because calcium binds primarily to elements of the C-terminal signature domain of thrombospondin-1, which are distant from the N-modules, such regulation was unexpected. To clarify the mechanism for this regulation, we compared ligand binding to the N-modules of thrombospondin-1 in the full-length protein and recombinant trimeric thrombospondin-1 truncated prior to the signature domain. Three monoclonal antibodies were identified that recognize the N-modules, two of which exhibit calcium-dependent binding to native thrombospondin-1 but not to the truncated trimeric protein. These antibodies or calcium selectively modulate interactions of fibronectin, heparin, sulfatide, alpha3beta1 integrin, tumor necrosis factor-alpha-stimulated gene-6 protein, and, to a lesser extent, alpha4beta1 integrin with native thrombospondin-1 but not with the truncated protein. These results indicate connectivity between calcium binding sites in the C-terminal signature domain and the N-modules of thrombospondin-1 that regulates ligand binding and functional activities of the N-modules.
通过去除钙而在血小板反应蛋白-1中诱导的构象变化调节其N模块与某些配体的相互作用。由于钙主要结合到血小板反应蛋白-1的C端特征结构域的元件上，而这些元件与N模块距离较远，所以这种调节是出乎意料的。为了阐明这种调节的机制，我们比较了全长蛋白中血小板反应蛋白-1的N模块与在特征结构域之前截短的重组三聚体血小板反应蛋白-1与配体的结合情况。鉴定出三种识别N模块的单克隆抗体，其中两种对天然血小板反应蛋白-1表现出钙依赖性结合，但对截短的三聚体蛋白则不表现。这些抗体或钙选择性地调节纤连蛋白、肝素、硫苷脂、α3β1整合素、肿瘤坏死因子-α刺激基因-6蛋白以及在较小程度上α4β1整合素与天然血小板反应蛋白-1的相互作用，但不调节与截短蛋白的相互作用。这些结果表明血小板反应蛋白-1的C端特征结构域中的钙结合位点与N模块之间存在联系，这种联系调节了N模块的配体结合和功能活性。
