Carlsson Karin, Osterlund Maria, Persson Egon, Freskgård Per-Ola, Carlsson Uno, Svensson Magdalena
IFM-Department of Chemistry, Linköping University, Sweden.
Biochim Biophys Acta. 2003 May 30;1648(1-2):12-6. doi: 10.1016/s1570-9639(03)00025-6.
We have used the site-directed labeling approach to study the Ca(2+)-dependent docking of factor VIIa (FVIIa) to soluble tissue factor (sTF). Nine Ca(2+) binding sites are located in FVIIa and even though their contribution to the overall binding between TF and FVIIa has been thoroughly studied, their importance for local protein-protein interactions within the complex has not been determined. Specifically we have monitored the association of the gamma-carboxyglutamic acid (Gla), the first EGF-like (EGF1), and the protease domains (PD) of FVIIa to sTF. Our results revealed that complex formation between sTF and FVIIa during Ca(2+) titration is initiated upon Ca(2+) binding to EGF1, the domain containing the site of highest Ca(2+) affinity. Besides we showed that a Ca(2+)-loaded Gla domain is required for an optimal association of all domains of FVIIa to sTF. Ca(2+) binding to the PD seems to be of some importance for the docking of this domain to sTF.
我们采用定点标记方法来研究因子VIIa（FVIIa）与可溶性组织因子（sTF）的钙依赖性对接。九个钙离子结合位点位于FVIIa中，尽管它们对TF和FVIIa之间整体结合的贡献已得到充分研究，但它们对复合物内局部蛋白质 - 蛋白质相互作用的重要性尚未确定。具体而言，我们监测了FVIIa的γ-羧基谷氨酸（Gla）、首个表皮生长因子样结构域（EGF1）和蛋白酶结构域（PD）与sTF的结合。我们的结果表明，在钙离子滴定过程中，sTF与FVIIa之间的复合物形成是在钙离子与EGF1结合后启动的，EGF1是含有最高钙离子亲和力位点的结构域。此外，我们还表明，钙离子负载的Gla结构域是FVIIa所有结构域与sTF实现最佳结合所必需的。钙离子与PD的结合似乎对该结构域与sTF的对接具有一定重要性。
