Proteolytic requirements for thrombin activation of anti-hemophilic factor (factor VIII).
Proteolytic requirements for thrombin activation of anti-hemophilic factor (factor VIII).
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抗血友病因子（因子 VIII）的凝血酶激活的蛋白水解要求。
DOI：
10.1073/pnas.85.8.2429
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发表时间：
1988
影响因子：
11.1
通讯作者：
Kaufman,RJ
中科院分区：
文献类型：
--
作者：
Pittman,DD;Kaufman,RJ
因子 VIII 在凝血的内在途径中发挥作用，作为因子 IXa 蛋白水解激活因子 X 的辅因子。蛋白水解裂解是激活所必需的，并且可能是辅因子活性失活的原因。为了确定因子 VIII 的激活和失活需要哪种多重切割，对因子 VIII cDNA 进行定点 DNA 介导的诱变，并在 COS-1 猴细胞中表达因子 VIII 的改变形式并进行表征。在740、1648和1721位裂解位点的氨基末端将精氨酸残基转化为异亮氨酸残基，导致修饰位点处的裂解抗性，但体外促凝血活性和对凝血酶激活的敏感性没有改变。对第 372 位或第 1689 位的凝血酶裂解位点进行类似修饰，产生具有残余因子 VIII 活性但在修饰位点抵抗凝血酶裂解且不易受凝血酶活化影响的分子。将残基 336 处的精氨酸修饰为异亮氨酸或赖氨酸，该位点被认为是被活化的蛋白 C 蛋白水解失活的位点，导致因子 VIII 分子的促凝血活性增加。这种活性的增加可能是由于对蛋白水解失活的更大抵抗力所致。提出了因子 VIII 激活和失活的模型。
Factor VIII functions in the intrinsic pathway of coagulation as the cofactor for factor IXa proteolytic activation of factor X. Proteolytic cleavage is required for activation and may be responsible for inactivation of cofactor activity. To identify which of the multiple cleavages are required for activation and inactivation of factor VIII, site-directed DNA-mediated mutagenesis of the factor VIII cDNA was performed and the altered forms of factor VIII were expressed in COS-1 monkey cells and characterized. Conversion of arginine residues to isoleucine residues at the aminoterminal side of the cleavage sites at positions 740, 1648, and 1721 resulted in cleavage resistance at the modified site with no alteration in the in vitro procoagulant activity and the susceptibility to thrombin activation. Similar modification of the thrombin cleavage sites at either position 372 or position 1689 resulted in molecules with residual factor VIII activity but resistant to thrombin cleavage at the modified site and not susceptible to thrombin activation. Modification of the arginine to either an isoleucine or a lysine at residue 336, the site postulated for proteolytic inactivation by activated protein C, resulted in a factor VIII molecule with increased procoagulant activity. This increased activity may result from greater resistance to proteolytic inactivation. A model for the activation and inactivation of factor VIII is proposed.