Cao Wenjing, Krishnaswamy Sriram, Camire Rodney M, Lenting Peter J, Zheng X Long
Departments of Pathology and Laboratory Medicine, Children's Hospital of Philadelphia, University of Pennsylvania, Philadelphia, PA 19104, USA.
Proc Natl Acad Sci U S A. 2008 May 27;105(21):7416-21. doi: 10.1073/pnas.0801735105. Epub 2008 May 20.
Proteolytic processing of von Willebrand factor (VWF) by ADAMTS13 metalloproteinase is crucial for normal hemostasis. In vitro, cleavage of VWF by ADAMTS13 is slow even at high shear stress and is typically studied in the presence of denaturants. We now show that, under shear stress and at physiological pH and ionic strength, coagulation factor VIII (FVIII) accelerates, by a factor of approximately 10, the rate of specific cleavage at the Tyr(1605)-Met(1606) bond in VWF. Multimer analysis reveals that FVIII preferentially accelerates the cleavage of high-molecular-weight multimers. This rate enhancement is not observed with VWF predenatured with 1.5 M guanidine. The ability of FVIII to enhance VWF cleavage by ADAMTS13 is rapidly lost after pretreatment of FVIII with thrombin. A FVIII derivative lacking most of the B domain behaves equivalently to full-length FVIII. In contrast, a derivative lacking both the B domain and the acidic region a3 that contributes to the high-affinity interaction of FVIII with VWF exhibits a greatly reduced ability to enhance VWF cleavage. Our data suggest that FVIII plays a role in regulating proteolytic processing of VWF by ADAMTS13 under shear stress, which depends on the high-affinity interaction between FVIII and its carrier protein, VWF.
血管性血友病因子（VWF）经ADAMTS13金属蛋白酶进行蛋白水解加工对于正常止血至关重要。在体外，即使在高剪切应力下，ADAMTS13对VWF的切割也很缓慢，并且通常在变性剂存在的情况下进行研究。我们现在表明，在剪切应力以及生理pH和离子强度条件下，凝血因子VIII（FVIII）使VWF中Tyr(1605)-Met(1606)键处的特异性切割速率加快了约10倍。多聚体分析显示，FVIII优先加速高分子量多聚体的切割。用1.5 M胍预变性的VWF未观察到这种速率增强。用凝血酶预处理FVIII后，FVIII增强ADAMTS13对VWF切割的能力迅速丧失。一种缺失大部分B结构域的FVIII衍生物的行为与全长FVIII相当。相反，一种既缺失B结构域又缺失有助于FVIII与VWF高亲和力相互作用的酸性区域a3的衍生物，增强VWF切割的能力大大降低。我们的数据表明，FVIII在剪切应力下通过ADAMTS13调节VWF的蛋白水解加工过程中发挥作用，这取决于FVIII与其载体蛋白VWF之间的高亲和力相互作用。
