Evidence for activation of tissue factor by an allosteric disulfide bond
Evidence for activation of tissue factor by an allosteric disulfide bond
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DOI：
10.1021/bi061271a
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发表时间：
2006-10-03
期刊：
影响因子：
2.9
通讯作者：
Hogg, Philip J.
中科院分区：
文献类型：
--
作者：
Chen, Vivien M.;Ahamed, Jasimuddin;Hogg, Philip J.
组织因子（TF）是哺乳动物质膜辅助因子，负责启动血液凝固。凝血因子 VIIa 与 TF 的结合通过有限的蛋白水解激活丝氨酸蛋白酶酶原因子 IX 和 X，从而形成稳定血栓的凝血酶和纤维蛋白网状结构。细胞质膜上的转录因子主要以隐秘构型存在，在受到某些刺激时会迅速转变为活性构型。 TF 的细胞外部分由两个 III 型纤连蛋白结构域组成。近膜结构域 (Cys186-Cys209) 中的二硫键对于此类结构域来说是非典型的，因为它连接同一片材中的相邻链，我们称之为变构键。通过突变两个半胱氨酸残基来消除变构二硫键会严重损害促凝血活性。硫醇烷基化剂 N-乙基马来酰亚胺和甲硫醇磺酸甲酯可阻断离子霉素对 TF 的活化，而硫醇氧化剂 HgCl2 和二硫醇交联剂可促进活化。 TF 激活不能用质膜外叶上磷脂酰丝氨酸的暴露来解释。隐秘 TF 含有未配对的半胱氨酸硫醇，这些半胱氨酸硫醇在激活后就会耗尽，并且解密与膜近端结构域的构象变化有关。这些发现意味着 Cys186-Cys209 二硫键在 TF 的隐秘形式中被还原，并且激活涉及二硫键的形成。该二硫键的形成可能会改变促进因子 IX 和 X 有效结合的结构域构象。
Tissue Factor (TF) is the mammalian plasma membrane cofactor responsible for initiation of blood coagulation. Binding of blood coagulation factor VIIa to TF activates the serine proteinase zymogens factors IX and X by limited proteolysis leading to the formation of a thrombin and fibrin meshwork that stabilizes the thrombus. TF on the plasma membrane of cells resides mostly in a cryptic configuration, which rapidly transforms into an active configuration in response to certain stimuli. The extracellular part of TF consists of two fibronectin type III domains. The disulfide bond in the membrane proximal domain (Cys186-Cys209) is atypical for domains of this type in that it links adjacent strands in the same, sheet, what we have called an allosteric bond. Ablation of the allosteric disulfide by mutating both cysteine residues severely impairs procoagulant activity. The thiol-alkylating agents N-ethylmaleimide and methyl methanethiolsulfonate block TF activation by ionomycin, while the thiol-oxidizing agent HgCl2 and dithiol cross-linkers promote activation. TF activation could not be explained by exposure of phosphatidylserine on the outer leaflet of the plasma membrane. Cryptic TF contained unpaired cysteine thiols that were depleted upon activation, and de-encryption was associated with a change in the conformation of the membrane-proximal domain. These findings imply that the Cys186-Cys209 disulfide bond is reduced in the cryptic form of TF and that activation involves formation of the disulfide. It is likely that formation of this disulfide bond changes the conformation of the domain that facilitates productive binding of factors IX and X.