Studies on Molecular Abnormality of Blood Coagulation and Fibrinolytic Factors

(1)两<alpha>种异常纤维蛋白原Kawaguchi和Osaka中精氨酸-16被半胱氨酸取代引起的纤维蛋白肽A释放缺陷：两种异常纤维蛋白原的先证者，命名为纤维蛋白原川口和纤维蛋白原Osaka，都是以纤维蛋白肽A（FPA）释放缺陷为特征的纤维蛋白原异常的杂合子。我们采用一种新的策略，利用溶无色杆菌M497-1的一种丝氨酸蛋白酶赖氨酰内肽酶，<alpha>从S-吡啶基乙基化A链的末端分离出异常的A1 -29肽<alpha>，并鉴定了<alpha>异常纤维蛋白原川口和大坂的AArg-16被Cys取代。(2)德岛岛凝血酶原：色氨酸替代精氨酸-418影响衍生凝血酶的纤维蛋白原凝血活性对一种遗传性异常的凝血酶原德岛进行了结构研究，以确定其在转化为凝血酶时纤维蛋白原凝血活性降低的原因。所用的凝血酶原样本来自杂合子，但仅含有缺陷的凝血酶原分子，因为患者是异常凝血酶原血症和低凝血酶原血症的杂合子。从异常凝血酶的赖氨酰内肽酶消化物中分离的肽的氨基酸序列分析表明，Arg-418（相当于胰凝乳蛋白酶编号系统中的Asn-101）已被Trp取代。这种氨基酸取代可以由Arg-418密码子中的单个核苷酸变化（CGG-&gt; TGG）引起。在凝血酶原德岛的凝血酶部分中发现的Arg-&gt; Trp置换似乎减少了其与包括纤维蛋白原和血小板受体在内的各种底物的相互作用，并解释了在先证者中观察到的复发性出血事件。

(1) DEFECTIVE RELEASE OF FIBRINOPEPTIDE A DUE TO SUBSTITUTION OF A <alpha> ARGININE-16 BY CYSTEINE IN TWO ABNORMAL FIBRINOGENS KAWAGUCHI AND OSAKA: The propositi of two abnormal fibrinogens, designated as fibrinogen Kawaguchi and fibrinogen Osaka, are both heterozygotes for the abnormality of fibrinogen characterized by defective release of fibrinopeptide A (FPA). By employing a new strategy of utilizing lysyl endopeptidase, a seryl protease derived from Achromobacter lyticus M497-1, we separated aberrant A <alpha> 1-29 peptides from the digests of the S-pyridylethylated A <alpha> chain and identified substituion of A <alpha> Arg-16 by Cys in both abnormal fibrinogens Kawaguchi and Osaka.(2) PROTHROMBIN TOKUSHIMA: A REPLACEMENT OF ARGININE-418 BY TRYPTOPHAN THAT IMPAIRS THE FIBRINOGEN CLOTTING ACTIVITY OF DERIVED THROMBIN TOKUSHIMA; Structural studies on a hereditarily abnormal prothrombin, prothrombin Tokushima, have been performed to identify the difference responsible for its reduced fibrinogen clotting activity upon conversion to thrombin. The prothrombin sample used was from a heterozygote but contained exclusively a defective prothrombin molecule, since the patient was heterozygous for both dysprothrombinemia and hypoprothrombinemia. Amino acid sequence analysis of a peptide isolated from a lysyl endopeptidase digest of the abnormal thrombin indicated that Arg-418 (equivalent to Asn-101 in the chymotrypsin numbering system) had been replaced by Trp. This amino acid substitution can result from a single nucleotide change in the codon for Arg-418 (CGG-> TGG). The Arg-> Trp replacement found in the thrombin portion of prothrombin Tokushima appears to reduce its interaction with various substrates including fibrinogen and platelet receptors, and accounts for the recurrent bleeding episode observed in the propositus.