Molecular and functional characterization of a natural homozygous Arg67His mutation in the prothrombin gene of a patient with a severe procoagulant defect contrasting with a mild hemorrhagic phenotype
Molecular and functional characterization of a natural homozygous Arg67His mutation in the prothrombin gene of a patient with a severe procoagulant defect contrasting with a mild hemorrhagic phenotype
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DOI：
10.1182/blood-2002-01-0243
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发表时间：
2002-08-15
期刊：
影响因子：
20.3
通讯作者：
Mannucci, PM
中科院分区：
文献类型：
--
作者：
Akhavan, S;De Cristofaro, R;Mannucci, PM
1例患者出现严重的血浆凝血功能缺陷，与非常轻微的出血性素质形成对照，在凝血酶原基因中发现纯合子Arg671His突变。野生型(FIIa[FIIa]-WT)和突变型Arg67His凝血酶(FIIa-MT67)具有相似的氨解活性。FIIa-WT和FIIa-MT67对纤维蛋白肽A的K(CAT)/K-m值分别为2.1×10(7)M(-1)S(-1)和9×10(5)M(-1)S(-1)。在FIIa-MT67和FIIa-WT中，蛋白C(PC)的活性降低与与血栓调节蛋白的结合亲和力降低33倍相关(TM；K-d=65.3 nM vs2.1 nM)。相反，在没有TM的情况下，PC的水解是正常的。Arg67His突变对蛋白酶激活的G蛋白偶联受体1(PAR-1)38-60肽的裂解有显著影响(k(CAT)/K-m=4×107M(-1)S(-1)至1.2×10(6)M(-1)S(-1))。FliA-MT67表现出较弱的血小板活化能力，这归因于PAR-1相互作用的缺陷，而与糖蛋白Ib的相互作用是正常的。与FIIa-WT相比，FIIa-MT67的与肝素辅因子11(HCII)相互作用的二级速率常数显著降低(高达500倍)，特别是在硫酸皮肤素存在的情况下，这表明在体内HCII对凝血酶的抑制作用受到严重损害。最后，Arg67His突变与凝血因子Xa-凝血因子Va复合体使凝血酶原激活降低5倍有关，可能是通过损害凝血酶原-凝血因子Va相互作用。这些实验表明，Arg67His替换对凝血酶的促凝和抗凝功能以及HCII对凝血酶的抑制作用都有显著的影响。轻度出血表型可以通过最终相互抵消的异常来解释。(C)2002年，由美国血液病学会公布。
In a patient who presented with a severe coagulation deficiency in plasma contrasting with a very mild hemorrhagic diathesis a homozygous Arg671His mutation was identified in the prothrombin gene. Wildtype (factor IIa [FIIa]-WT) and mutant Arg67His thrombin (FIIa-MT67) had similar amidolytic activity. By contrast, the k(cat)/K-m value of fibrinopeptide A hydrolysis by FIIa-WT and FIIa-MT67 was equal to 2.1 X 10(7) M(-1)s(-1) and 9 x 10(5) M(-1)s(-1). Decreased activation of protein C (PC) correlated with the 33-fold decreased binding affinity for thrombomodulin (TM; K-d = 65.3 nM vs 2.1 nM, in FIIa-MT67 and in FIIa-WT, respectively). In contrast, hydrolysis of PC in the absence of TM was normal. The Arg67His mutation had a dramatic effect on the cleavage of protease-activated G protein-coupled receptor 1 (PAR-1) 38-60 peptide (k(cat)/K-m = 4 x 107 M(-1)s(-1) to 1.2 x 10(6) M(-1)s(-1)). Flia-MT67 showed a weaker platelet activating capacity, attributed to a defective PAR-1 interaction, whereas the interaction with glycoprotein Ib was normal. A drastic decrease (up to 500-fold) of the second-order rate constant pertaining to heparin cofactor 11 (HCII) interaction, especially in the presence of dermatan sulfate, was found for the FIIa-MT67 compared with FIIa-WT, suggesting a severe impairment of thrombin inhibition by HCII in vivo. Finally, the Arg67His mutation was associated with a 5-fold decrease of prothrombin activation by the factor Xa-factor Va complex, perhaps through impairment of the prothrombin-factor Va interaction. These experiments show that the Arg67His substitution affects drastically both the procoagulant and the anticoagulant functions of thrombin as well as its inhibition by HCII. The mild hemorrhagic phenotype might be explained by abnormalities that ultimately counterbalance each other. (C) 2002 by The American Society of Hematology.