Studies on Molecular Abnormality of Blood Coagulation and Fibrinolytic Factors

(1)两种异常纤维蛋白原中的A < α >精氨酸-16被半胱氨酸取代导致纤维蛋白肽A释放缺陷：两种异常纤维蛋白原的阳性，被称为纤维蛋白原KAWAGUCHI和纤维蛋白原OSAKA，都是纤维蛋白原异常的杂合子，其特征是纤维蛋白肽A （FPA）释放缺陷。利用lysyl内肽酶（一种源自lyacromobacter lyticus M497-1的丝氨酸蛋白酶）的新策略，我们从s - pyrydylethylated a < α >链的消化液中分离出异常的a < α > 1-29肽，并在异常纤维蛋白原Kawaguchi和Osaka中鉴定出a < α > Arg-16被Cys取代。(2)德岛凝血酶原：色氨酸替代精氨酸-418，损害衍生的德岛凝血酶的纤维蛋白原凝血活性；对一种遗传性异常凝血酶原，即德岛凝血酶原进行了结构研究，以确定其转化为凝血酶时纤维蛋白原凝血活性降低的差异。使用的凝血酶原样本来自杂合子，但只含有有缺陷的凝血酶原分子，因为患者是凝血酶原异常和凝血酶原低的杂合子。从异常凝血酶的赖氨酸内肽酶消化中分离的肽的氨基酸序列分析表明，Arg-418（相当于凝乳胰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 （CGG-> TGG）密码子的单个核苷酸变化引起。在德岛凝血酶原凝血酶部分发现的Arg->色氨酸替代似乎减少了其与各种底物（包括纤维蛋白原和血小板受体）的相互作用，并解释了在患者中观察到的反复出血事件。

(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digest of the abnormal thrombin indicated that Arg-418 (equivalent to Asn-101 in the chymotrypsin numbering system) had been replaced by Trp.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.