Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.5 (thrombin)
33,306 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Coagulation factor V (fV) is a single-chain glycoprotein (Mr 330,000; domain structure A1-A2-B-A3-C1-C2) that is activated to factor Va (fVa; Mr 174,000) by thrombin, which cleaves away the B domain leaving a heterodimeric structure composed of a heavy chain (A1-A2; Mr 94,000) and a light chain (A3-C1-C2; Mr 74,000). We analyzed the ultrastructure of scanning transmission electron microscope images of bovine and human fV, bovine fVa, and its constituent light chains and heavy chains. Factor V molecules had irregularly globular (10-12 nm) to oblong (8-14 nm) core structures which commonly displayed a peripheral satellite appendage of variable morphology attached to the core by a narrow stalk. Scanning transmission electron microscope mass analyses indicated that monomolecular bovine fV molecules had a mass of 322 +/- 45 kDa and human fV, 315 +/- 31 kDa. Factor Va molecules were irregular, globular (8-12 nm) structures that resembled the fV core structure, lacked the satellite appendage representing B domainal structures, and had a mass of 180 +/- 22 kDa. Our findings permit us to propose a structural model of fV suggesting the relative orientation of its closely associated light chain and heavy chain core components and indicating that these constituents remain associated in the transition from fV to fVa.
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PMID:Structural model of factors V and Va based on scanning transmission electron microscope images and mass analysis. 234 7

Coagulation factor V is a high molecular weight plasma glycoprotein that participates as a cofactor in the conversion of prothrombin to thrombin by factor Xa. A phage lambda gt11 Hep G2 cell cDNA expression library was screened by using an affinity-purified antibody to human factor V, and 11 positive clones were isolated and plaque-purified. The clone containing the largest cDNA insert contained 2970 nucleotides and coded for 938 amino acids, a stop codon, and 155 nucleotides of 3' noncoding sequence including a poly(A) tail. The coding region includes 651 amino acids from the carboxyl terminus that constitute the light chain of human factor Va and 287 amino acids that are part of the connecting region of the protein. The predicted amino acid sequence agreed completely with 147 amino acid residues that were identified by Edman degradation of cyanogen bromide peptides isolated from the light chain. During the activation of factor V, several peptide bonds are cleaved by thrombin, giving rise to a heavy chain, a connecting fragment(s), and a light chain. The light chain is generated by the cleavage of an Arg-Ser peptide bond. The amino acid sequence of the light chain is homologous (40%) with the carboxyl-terminal fragment (Mr, 73,000) of human factor VIII. Both fragments have a similar domain structure that includes a single ceruloplasmin-related domain followed by two C domains. The carboxyl terminus of the connecting region, however, shows no significant amino acid sequence homology with factor VIII. It is very acidic and contains a number of potential N-linked glycosylation sites. It also contains about 20 tandem repeats of nine amino acids.
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PMID:Cloning of a cDNA coding for human factor V, a blood coagulation factor homologous to factor VIII and ceruloplasmin. 309 20

Coagulation factor V is a critical cofactor for the activation of prothrombin to thrombin, the penultimate step in the generation of a fibrin blood clot. Genetic deficiency of factor V results in a congenital bleeding disorder (parahaemophilia), whereas inheritance of a mutation rendering factor V resistant to inactivation is an important risk factor for thrombosis. We report here that approximately half of homozygous embryos deficient in factor V (Fv-/-), which have been generated by gene targeting, die at embryonic day (E) 9-10, possibly as a result of an abnormality in the yolk-sac vasculature. The remaining Fv-/- mice progress normally to term, but die from massive haemorrhage within 2 hours of birth. Considered together with the milder phenotypes generally associated with deficiencies of other clotting factors, our findings demonstrate the primary role of the common coagulation pathway and the absolute requirement for functional factor V for prothrombinase activity. They also provide direct evidence for the existence of other critical haemostatic functions for thrombin in addition to fibrin clot formation, and identify a previously unrecognized role for the coagulation system in early mammalian development.
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PMID:Fatal haemorrhage and incomplete block to embryogenesis in mice lacking coagulation factor V. 890 Feb 78

Coagulation factor V circulates in plasma as a single chain protein which expresses little procoagulant activity. After its activation by limited proteolysis by thrombin or factor Xa, factor Va functions as cofactor to factor Xa in the activation of prothrombin. Thrombin cleaves human factor V at Arg709, Arg1018 and Arg1545 and factor Va is formed by the heavy and light chains, which correspond to the N-terminal and C-terminal fragments, respectively. Factor Xa has been shown to cleave factor V at Arg1018 and at a second undefined position close to Arg709. The factor-Xa-mediated cleavage at Arg1018 has been proposed to be sufficient for expression of full factor Va activity. To study the activation of factor V by factor Xa, site-directed mutagenesis was used to convert Arg709 to Gln, Arg1018 to Ile, and Arg1545 to Gln. Constructs containing all possible combinations of native and mutated residues in these positions were expressed transiently in COS 1 cells. The various factor-V mutants were incubated with factor Xa or thrombin. The proteolytic cleavage pattern was analyzed by Western blotting, and the specific factor-Va activities determined in a prothrombinase assay. Control experiments using thrombin gave results which were in agreement with those on record, i.e. cleavages at both Arg709 and Arg1545 were required for expression of full factor-Va activity, whereas the cleavage at Arg1018 enhanced the rate of cleavage at Arg1545. Factor Xa was found to cleave factor V at all three thrombin cleavage sites, i.e. at Arg709, Arg1018 and Arg1545. An additional factor-Xa-cleavage site was found in the light chain region at Arg1765. Cleavage at Arg1018 by factor Xa was not sufficient for expression of full factor-Va activity. Full factor-Va activity was only obtained after cleavage at both Arg709 and Arg1545. The factor-Xa-mediated cleavage at Arg709 was kinetically favourable over that at Arg1545. Factor V which was mutated at all three sites (at positions 709, 1018 and 1545) was resistant to activation by thrombin. However, treatment with factor Xa yielded an increased factor-Va activity which was associated with the cleavage at Arg1765. Our study extends previously results on thrombin activation of factor V and elucidates the relative importance of the different cleavage sites for activation of factor V by factor Xa.
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PMID:Cleavage requirements for activation of factor V by factor Xa. 924 3

Coagulation factor V is composed of domains A1-A2-B-A3-C1-C2 and is activated by thrombin through proteolytic cleavage at Arg 709, Arg 1018 and Arg 1545. Upon thrombin activation, the B-domain is released and the active factor Va is formed by the heavy (A1-A2) and light chains (A3-C1-C2). Factor Va functions as an essential cofactor to factor Xa in the conversion of prothrombin to thrombin during coagulation. Recently it was shown that coagulation factor V, apart from being a precursor form to the procoagulant factor Va, also has anticoagulant properties, as it functions as a cofactor to activated protein C (APC). APC is a member of the anticoagulant pathway and downregulates the coagulation process through proteolytic inactivation of factors VIII/VIIIa and factors V/Va. In a factor VIIIa degradation assay, the APC-mediated inactivation of factor VIIIa is potentiated by the synergistic cofactors protein S and factor V. Protein S alone has little cofactor activity, whereas in the presence of factor V it is dramatically enhanced. This study provides insights into the molecular mechanisms that regulate the anticoagulant activity of factor V. Thrombin cleavage of factor V occurs in a sequential order. The thrombin cleavage site Arg 1545 is kinetically less favored than the other two sites, and cleavage at this site is the last to occur during thrombin activation of factor V As a consequence of this, different activation intermediates exist that express different levels of procoagulant activity. The anticoagulant activities of these intermediates have now been studied. It was found that factor V could be cleaved by thrombin at both Arg 709 and Arg 1018 and still work fully as a cofactor to APC, whereas cleavage at Arg 1545 completely abolished the anticoagulant activity of factor V. This suggests that the APC cofactor function of factor V depends on the B-domain remaining attached to the A3 domain. This study further shows that APC converts coagulation factor V into a member of the anticoagulant pathway by cleaving factor V in the A2 domain at Arg 506. By cleavage of factor V, APC not only produces an anticoagulant cofactor, but at the same time eliminates the pool of procoagulant factor V, since APC cleaved factor V will have no future as a cofactor in the coagulation. The unique way by which APC and thrombin, through proteolytic cleavage, can convert factor V into either an anticoagulant or a procoagulant adds to the intriguing mechanisms that balance the procoagulant and anticoagulant forces.
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PMID:Mechanisms that regulate the anticoagulant function of coagulation factor V. 1009 86

Coagulation factor V (FV) circulates in two forms, FV1 and FV2, having slightly different molecular masses and phospholipid-binding properties. The aim was to determine whether this heterogeneity is due to the degree of glycosylation of Asn(2181). FVa1 and FVa2 were isolated and digested with endoglycosidase PNGase F. As judged by Western blotting, the FVa2 light chain contained two N-linked carbohydrates, whereas FVa1 contained three. Wild-type FV and three mutants, Asn(2181)Gln, Ser(2183)Thr, and Ser(2183)Ala, were expressed in COS1 cells, activated by thrombin, and analyzed by Western blotting. Wild-type FVa contained the 71 kDa-74 kDa doublet, whereas the Asn(2181)Gln and Ser(2183)Ala mutants contained only the 71 kDa light chain. In contrast, the Ser(2183)Thr mutant gave a 74 kDa light chain. This demonstrated that the third position in the Asn-X-Ser/Thr consensus affects glycosylation efficiency, Thr being associated with a higher degree of glycosylation than Ser. The Ser(2183)Thr mutant FVa was functionally indistinguishable from plasma-purified FVa1, whereas Asn(2181)Gln and Ser(2183)Ala mutants behaved like FVa2. Thus, the carbohydrate at Asn(2181) impaired the interaction between FVa and the phospholipid membrane, an interpretation consistent with a structural analysis of a three-dimensional model of the C2 domain and the position of a proposed phospholipid-binding site. In conclusion, we show that the FV1-FV2 heterogeneity is caused by differential glycosylation of Asn(2181) related to the presence of a Ser rather than a Thr at the third position in the consensus sequence of glycosylation.
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PMID:Partial glycosylation of Asn2181 in human factor V as a cause of molecular and functional heterogeneity. Modulation of glycosylation efficiency by mutagenesis of the consensus sequence for N-linked glycosylation. 1052 Dec 65

Coagulation factor V (FV) plays an important role in maintaining the hemostatic balance in both the formation of thrombin in the procoagulant pathway as well as in the protein C anticoagulant pathway. FV deficiency is a rare bleeding disorder with variable phenotypic expression. Little is known about the molecular basis underlying this disease. This study identified 5 novel mutations associated with FV deficiency in 3 patients with severe FV deficiency but different clinical expression and 2 unaffected carriers. Four mutations led to a premature termination codon either by a nonsense mutation (single-letter amino acid codes): A1102T, K310Term. (FV Amersfoort) and C2491T, Q773Term. (FV Casablanca) or a frameshift: an 8-base pair deletion between nucleotides 1130 and 1139 (FV Seoul(1)) and a 1-base pair deletion between nucleotides 4291 and 4294 (FV Utrecht). One mutation was a novel missense mutation: T1927C, C585R (FV Nijkerk), resulting in the absence of mutant protein despite normal transcription to RNA. Most likely, an arginine at this position disrupts the hydrophobic interior of the FV A2 domain. The sixth detected mutation was a previously reported missense mutation: A5279G, Y1702C (FV Seoul(2)). In all cases, the presence of the mutation was associated with type I FV deficiency. Identifying the molecular basis of mutations underlying this rare coagulation disorder will help to obtain more insight into the mechanisms involved in the variable clinical phenotype of patients with FV deficiency.
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PMID:Five novel mutations in the gene for human blood coagulation factor V associated with type I factor V deficiency. 1143 4

Coagulation factor V (FV) is the protein cofactor required in vivo for the rapid generation of thrombin catalyzed by the prothrombinase complex. It also represents a central regulator in the early phases of blood clot formation, as it contributes to the anticoagulant pathway by participating in the downregulation of factor VIII activity. Conversion of precursor FV to either a procoagulant or anticoagulant cofactor depends on the local concentration of procoagulant and anticoagulant enzymes, so that FV may be regarded as a daring tight-rope walker gently balancing opposite forces. Given this dual role, genetic defects in the FV gene may result in opposite phenotypes (hemorrhagic or thrombotic). Besides a concise description on the structural, procoagulant and anticoagulant properties of FV, this review will focus on bleeding disorders associated with altered levels of this molecule. Particular attention will be paid to the mutational spectrum of type I FV deficiency, which is characterized by a remarkable genetic heterogeneity and by an uneven distribution of mutations throughout the FV gene.
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PMID:Inherited defects of coagulation factor V: the hemorrhagic side. 1705 27

Coagulation factor V (FV) is an essential component of the prothrombinase complex, which activates the zymogen prothrombin to thrombin. Acquired FV inhibitor is rare and clinical symptoms are quite variable. The aim of this study was to summarize the spectrum of the bleeding presentation of acquired FV deficiency and characterize the underlying causes of the clinical symptoms. This study was designed as a descriptive retrospective and 30 case reports were included for further analysis. At least 33 cases of acquired FV inhibitor were investigated. Most patients have a presentation of bleeding and most of those are from hematuria and bleeding at surgical sites. Seven cases of asymptomatic acquired FV inhibitor were also detected. A total of 39.4% (19 of 33) of those cases with acquired FV inhibitor have an unknown cause. For those with known etiologies, chemical and drug-induced causes are the most common (30.8%).
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PMID:Spectrum of bleeding in acquired factor V inhibitor: a summary of 33 cases. 1700 Aug 94

Coagulation factor V (FV), present in plasma and platelets, is an indispensable clotting factor, as demonstrated by the uniform lethality of FV knock-out mice. Surprisingly, however, severe FV deficiency is rarely fatal in humans. In fact, although several cases of life-threatening intracranial haemorrhage have been reported in FV-deficient newborns, many patients with undetectable FV levels experience only mild to moderate bleeding and do not require routine prophylaxis. While the reasons for this variable phenotypic expression are largely unknown, several observations from different laboratories indicate platelets as crucial players in FV deficiency. Moreover, we have recently shown that plasma levels of tissue factor pathway inhibitor are considerably reduced in FV-deficient plasma, which results in enhanced thrombin generation especially at very low FV levels (<2%). The present review discusses and integrates these findings in the context of the biology of FV and the clinical features of FV deficiency.
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PMID:Advances in understanding the bleeding diathesis in factor V deficiency. 1943 79


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