Zinc-dependent conformational changes in domain D5 of high molecular mass kininogen modulate contact activation
Zinc-dependent conformational changes in domain D5 of high molecular mass kininogen modulate contact activation
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DOI：
10.1046/j.1432-1033.2001.01888.x
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发表时间：
2001-01-01
期刊：
影响因子：
--
通讯作者：
Sjöbring, U
中科院分区：
文献类型：
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作者：
Herwald, H;Mörgelin, M;Sjöbring, U
人高分子量激肽原（HK）作为非酶辅助因子参与接触系统。在这里，我们表明，重组域D5的HK（rD5）缩短凝血的内源性途径的凝血时间，并减弱缓激肽的产生。进一步的研究表明，HK内D5结构域的正确折叠是激活接触系统所必需的。rD5的折叠似乎受到金属离子Zn 2+、Ni 2+和Cu 2+的调节，因为针对HK中锌结合位点的特异性抗体以金属离子浓度依赖性方式结合HK和rD5。这三种金属离子特别影响接触活化的发现表明，它们调节rD5对带负电荷表面的可及性。通过rD5的荧光光谱法获得了对所观察到的现象是由于构象变化的假设的支持，表明其荧光光谱在ZnCl2的存在下发生了变化。此外，负染电镜实验表明，锌诱导的D5的变化也影响整个HK蛋白的构象。目前的数据强调的作用，锌和其他金属离子的调节接触激活。
Human high molecular mass kininogen (HK) participates as nonenzymatic cofactor in the contact system. Here, we show that recombinant domain D5 of HK (rD5) prolongs the clotting time of the intrinsic pathway of coagulation and attenuates the generation of bradykinin. Further studies indicate that a correct fold of domain D5 within HK is required for the activation of the contact system. The folding of rD5 seems to be modulated by the metal ions Zn2+, Ni2+, and Cu2+ as a specific antibody directed against the zinc-binding site in HK binds to HK and rD5 in a metal ion concentration dependent manner. The finding that these three metal ions specifically affect contact activation suggests that they regulate the accessibility of rD5 for negatively charged surfaces. Support for the assumption that the observed phenomena are due to conformational changes was obtained by fluorescence spectroscopy of rD5, demonstrating that its fluorescence spectrum was changed in the presence of ZnCl2. Moreover, negative staining electron microscopy experiments suggest that the zinc-induced changes in D5 also affect the conformation of the entire HK protein. The present data emphasize the role of zinc and other metal ions in the regulation of contact activation.