Phosphatidylserine and phosphatidylethanolamine regulate the structure and function of FVIIa and its interaction with soluble tissue factor.
Phosphatidylserine and phosphatidylethanolamine regulate the structure and function of FVIIa and its interaction with soluble tissue factor.
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磷脂酰丝氨酸和磷脂酰乙醇胺调节FVIIA的结构和功能及其与可溶性组织因子的相互作用。
DOI：
10.1042/bsr20204077
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发表时间：
2021-02-26
影响因子：
4
通讯作者：
Majumder R
中科院分区：
文献类型：
--
作者：
Sengupta T;Koklic T;Lentz BR;Majumder R
细胞膜在凝血级联的许多步骤中具有重要功能，包括通过因子VIIa（FVIIa）-组织因子（TF）复合物（外源性Xase）激活因子X（FX）。FVIIa与因子IXA（FIXa）和FXA具有结构相似性。FIXa和FXa通过其γ-羧基谷氨酸富集结构域（Gla）和表皮生长因子（EGF）结构域与含磷脂酰丝氨酸（PS）的膜结合来调节。尽管FVIIa也具有富含Gla的区域，但其对含PS的膜的亲和力比FIXa和FXa低得多。研究表明，更常见的内皮细胞脂质磷脂酰乙醇胺（PE）可能会增加PS在FVIIa膜结合和蛋白水解活性中的作用。我们使用可溶形式的PS和PE（1，2-二己酰基-sn-甘油基-3-磷酸-1-丝氨酸（C6 PS）、1，2-二己酰基-sn-甘油基-3-磷酸-乙醇胺（C6 PE））来检验这两种脂质共同结合FVIIa以促进FVIIa膜结合和蛋白水解活性的假设。通过平衡透析和色氨酸荧光分析，我们发现FVIIa上有两个位点与C6 PE和C6 PS的结合力相等，Kd值为150-160 μM，但Gla结构域的缺失降低了结合力。当通过FVIIa蛋白水解活性监测时，脂质的结合具有更大的亲和力（Kd = 70-80 μM）。所有数据集的全局拟合表明每种脂质的两个分子的独立结合。在可溶性TF（sTF）加C6 PS/C6 PE的存在下，FVIIa的蛋白水解活性增加100 -100倍。然而，在sTF存在下，Gla缺失的FVIIa的蛋白水解活性急剧降低，这表明Gla结构域对于维持完整的蛋白水解活性的重要性。
Cell membranes have important functions in many steps of the blood coagulation cascade, including the activation of factor X (FX) by the factor VIIa (FVIIa)-tissue factor (TF) complex (extrinsic Xase). FVIIa shares structural similarity with factor IXa (FIXa) and FXa. FIXa and FXa are regulated by binding to phosphatidylserine (PS)-containing membranes via their γ-carboxyglutamic acid-rich domain (Gla) and epidermal growth-factor (EGF) domains. Although FVIIa also has a Gla-rich region, its affinity for PS-containing membranes is much lower compared with that of FIXa and FXa. Research suggests that a more common endothelial cell lipid, phosphatidylethanolamine (PE), might augment the contribution of PS in FVIIa membrane-binding and proteolytic activity. We used soluble forms of PS and PE (1,2-dicaproyl-sn-glycero-3-phospho-l-serine (C6PS), 1,2-dicaproyl-sn-glycero-3-phospho-ethanolamine (C6PE)) to test the hypothesis that the two lipids bind to FVIIa jointly to promote FVIIa membrane binding and proteolytic activity. By equilibrium dialysis and tryptophan fluorescence, we found two sites on FVIIa that bound equally to C6PE and C6PS with Kd of ∼ 150–160 μM, however, deletion of Gla domain reduced the binding affinity. Binding of lipids occurred with greater affinity (Kd∼70–80 μM) when monitored by FVIIa proteolytic activity. Global fitting of all datasets indicated independent binding of two molecules of each lipid. The proteolytic activity of FVIIa increased by ∼50–100-fold in the presence of soluble TF (sTF) plus C6PS/C6PE. However, the proteolytic activity of Gla-deleted FVIIa in the presence of sTF was reduced drastically, suggesting the importance of Gla domain to maintain full proteolytic activity.