scholarly journals A gamma Gly-268 to Glu substitution is responsible for impaired fibrin assembly in a homozygous dysfibrinogen Kurashiki I

Blood ◽  
1996 ◽  
Vol 87 (11) ◽  
pp. 4686-4694 ◽  
Author(s):  
K Niwa ◽  
M Takebe ◽  
T Sugo ◽  
Y Kawata ◽  
J Mimuro ◽  
...  
Keyword(s):  
Tissue Type ◽  
Factor Xiiia ◽  
Cross Linking ◽  
Fibrin Gel ◽  
Functional Abnormality ◽  
Human Fibrinogen ◽  
Fibrin Monomer ◽  
Longitudinal Association ◽  
Self Association ◽  
A Site

A new type of gamma Gly-268 (GGA) to Glu (GAA) substitution has been identified in a homozygous dysfibrinogen by analyses of the affected polypeptide and its encoding gene derived from a 58 year-old man manifesting no major bleeding or thrombosis. The functional abnormality was characterized by impaired fibrin assembly most likely due to failure to construct properly aligned double-stranded fibrin protofibrils. This presumption was deduced from the following findings: (1) Factor XIIIa-catalyzed cross-linking of the fibrin gamma-chains progressed in a normal fashion, indicating that the contact between the central E domain of one fibrin monomer and the D domain of another took place normally; (2) Nevertheless, factor XIIIa-catalyzed cross-linking of the fibrinogen gamma-chains was obviously delayed, suggesting that longitudinal association of D domains of different fibrin monomers, ie, D:D association was perturbed; (3) Plasminogen activation catalyzed by tissue-type plasminogen activator was not as efficiently facilitated by polymerizing fibrin monomer derived from the patient as by the normal counterpart. Therefore, gamma Gly-268 would not be involved in the 'a' site residing in the D domain, which functions as a complementary binding site with the thrombin-activated 'A' site in the central E domain, but would be rather involved in the D:D self association sites recently proposed for human fibrinogen. Thus, the gamma Glu-268 substitution newly identified in this homozygous dysfibrinogen seems to impair proper alignment of adjacent D domains of neighboring fibrin molecules in the double-stranded fibrin protofibril, resulting in delayed fibrin gel formation.

Myosin: a noncovalent stabilizer of fibrin in the process of clot dissolution

Blood ◽  
2003 ◽  
Vol 101 (11) ◽  
pp. 4380-4386 ◽  
Author(s):  
Krasimir Kolev ◽  
Kiril Tenekedjiev ◽  
Katalin Ajtai ◽  
Ilona Kovalszky ◽  
Judit Gombás ◽  
...  
Keyword(s):  
Plasminogen Activator ◽  
Rate Constant ◽  
Dissociation Constants ◽  
Dissociation Rate ◽  
Covalent Modification ◽  
Tissue Type ◽  
Factor Xiiia ◽  
Molar Ratio ◽  
Fibrin Monomer

Abstract Myosin modulates the fibrinolytic process as a cofactor of the tissue plasminogen activator and as a substrate of plasmin. We report now that myosin is present in arterial thrombi and it forms reversible noncovalent complexes with fibrinogen and fibrin with equilibrium dissociation constants in the micromolar range (1.70 and 0.94 μM, respectively). Competition studies using a peptide inhibitor of fibrin polymerization (glycl-prolyl-arginyl-proline [GPRP]) indicate that myosin interacts with domains common in fibrinogen and fibrin and this interaction is independent of the GPRP-binding polymerization site in the fibrinogen molecule. An association rate constant of 1.81 × 102 M–1 · s–1 and a dissociation rate constant of 3.07 × 10–4 s–1 are determined for the fibrinogen-myosin interaction. Surface plasmon resonance studies indicate that fibrin serves as a matrix core for myosin aggregation. The fibrin clots equilibrated with myosin are stabilized against dissolution initiated by plasminogen and tissue-type plasminogen activator (tPA) or urokinase (at fibrin monomer-myosin molar ratio as high as 30) and by plasmin under static and flow conditions (at fibrin monomer-myosin molar ratio lower than 15). Myosin exerts similar effects on the tPA-induced dissolution of blood plasma clots. Covalent modification involving factor XIIIa does not contribute to this stabilizing effect; myosin is not covalently attached to the clot by the time of complete cross-linking of fibrin. Thus, our in vitro data suggest that myosin detected in arterial thrombi binds to the polymerized fibrin, in the bound form its tPA-cofactor properties are masked, and the myosinfibrin clot is relatively resistant to plasmin.

Clottability and Cross-Linking Reactivity of Fibrin(ogen) Following Differential Release of Fibrinopeptides A and B

1976 ◽  
Vol 36 (03) ◽  
pp. 582-592 ◽  
Author(s):  
M Furlan ◽  
T Seelich ◽  
E. A Beck
Keyword(s):  
Early Stage ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Factor Xiiia ◽  
Cross Linking ◽  
Human Fibrinogen ◽  
Prolonged Incubation ◽  
Fibrin Networks ◽  
Covalently Linked ◽  
Bothrops Atrox

SummaryHuman fibrinogen was treated at pH 6.0, 7.3 and 9.0 with thrombin, batroxobin (thrombinlike fraction of Bothrops atrox venom) or an extract of the venom from Ancistrodon contortrix contortrix. These three enzymes released the NH2-terminal fibrinopeptides A and B at different rates. Thrombin-free, preactivated factor XIII (factor XIIIa) was added to incubation mixtures to stabilize resulting fibrin(ogen) aggregates. Cross-linking of γ-chains and the size of covalently linked fibrin-fibrinogen oligomers were studied in an early stage of fibrinopeptide cleavage using polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate.Batroxobin (pH 7.3) and thrombin (pH. 6.0) preferentially released fibrinopeptide A, and resulting fibrin aggregates became rapidly insoluble. However, when fibrinopeptide B was removed with the contortrix enzyme, soluble cross-linked oligomers appeared initially. The opaque fibrin clots, produced by thrombin (pH 6.0) or contortrix procoagulant fraction (pH 7.3), were found to be devoid of α-polymers even after prolonged incubation with factor XIIIa. Our data suggest that the solubility and opacity of fibrin networks are not primarily related to the type of the cross-link (γ-γ versus α-α interactions).

Factor XIIIa Cross-Linking of the Marburg Fibrin: Formation of αm·γn-Heteromultimers and the α-Chain–Linked Albumin·γ Complex, and Disturbed Protofibril Assembly Resulting in Acquisition of Plasmin Resistance Relevant to Thrombophila

Blood ◽  
1998 ◽  
Vol 91 (9) ◽  
pp. 3282-3288 ◽  
Author(s):  
Teruko Sugo ◽  
Chizuko Nakamikawa ◽  
Mikihiro Takebe ◽  
Isao Kohno ◽  
Rudorf Egbring ◽  
...  
Keyword(s):  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Caesarian Section ◽  
Tissue Type ◽  
Factor Xiiia ◽  
Cross Linking ◽  
Amino Acid Residues ◽  
Pelvic Vein ◽  
Vein Thrombosis ◽  
Type Plasminogen Activator

The truncated Aα-chain of fibrinogen Marburg is partly linked with albumin by a disulfide bond. Based on the recovery of the first six amino acid residues assigned to the subunit polypeptides of fibrinogen (the Aα-and γ-chains) and albumin, 0.33 mol of albumin was estimated to be linked to 1 mol of the Marburg fibrinogen. When the Marburg fibrinogen was clotted with thrombin-factor XIIIa-Ca2+, various αmγnheteromultimers were produced, and part of the albumin was cross-linked to the γ-chain. Acid-solubilized Marburg fibrin monomer failed to form large aggregates that could be detected by monitoring turbidity at A350, but it was able to enhance tissue-type plasminogen-activator–catalyzed plasmin generation, though not as avidly as the normal control, indicating that the double-stranded protofibrils had, to some extent, been constructed. This idea seems to be supported by normal factor XIIIa–catalyzed cross-linking of the fibrin γ-chains. However, the cross-linked Marburg fibrin, being apparently fragile and translucent, was highly resistant against plasmin, and its subunit components were considerably retained for 48 hours as noted by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Although the exact mechanisms are still unclear, the albumin-incorporated factor XIIIa–cross-linked Marburg fibrin seems to have undergone a critical structural alteration(s) to acquire resistance against plasmin. This aquisition of plasmin resistance may be contributed to the postoperative pelvic vein thrombosis and recurrent pulmonary embolisms in the patient after caesarian section for her first delivery at the age of 20 years.

Factor XIIIa-Catalyzed Cross-Linking of Recombinant αC Fragments of Human Fibrinogen†

Biochemistry ◽  
1996 ◽  
Vol 35 (18) ◽  
pp. 5810-5816 ◽  
Author(s):  
Yury V. Matsuka ◽  
Leonid V. Medved ◽  
Mary M. Migliorini ◽  
Kenneth C. Ingham
Keyword(s):  
Factor Xiiia ◽  
Cross Linking ◽  
Human Fibrinogen

A Hamster Antibody to the Mouse Fibrinogen Chain Inhibits Platelet-fibrinogen Interactions and FXIIIa-mediated Fibrin Cross-linking, and Facilitates Thrombolysis

2001 ◽  
Vol 86 (10) ◽  
pp. 1047-1056 ◽  
Author(s):  
Markéta JirouȈsková ◽  
Susan Smyth ◽  
Bohdan Kudryk ◽  
Barry Coller
Keyword(s):  
Factor Xiiia ◽  
Cross Linking ◽  
Murine Models ◽  
Clot Retraction ◽  
Human Fibrinogen ◽  
Hemostatic System ◽  
Fibrinogen Binding ◽  
C Terminus ◽  
New Information ◽  
Tissue Plasminogen

SummaryMurine models employing genetically altered mice have the potential to provide important new information about the hemostatic system, but before such data can be extrapolated to humans it is necessary to define the similarities and differences between murine and human hemostasis. After establishing the similarities of murine fibrinogen to human fibrinogen in its pattern of proteolysis in response to plasmin and its cross-linking by factor XIIIa, we studied a new hamster monoclonal antibody (mAb) 7E9 that reacts with the γ chain of mouse fibrinogen. This antibody inhibits platelet adhesion to fibrinogen, platelet-mediated clot retraction, platelet aggregation, and FXIIIa-mediated cross-linking of fibrin; it also facilitates tissue plasminogen activator (tPA)-mediated lysis of fibrin formed either in the absence or presence of platelets. These data provide evidence that the C-terminus of mouse fibrinogen γ chain, like that of human fibrinogen, is involved in fibrinogen binding to platelets and FXIIIa-mediated cross-linking of fibrin. Our data raise the possibility that a therapeutic agent that targets the C-terminus of the γ chain in human fibrinogen might have broad antithrombotic and profibrinolytic effects.

scholarly journals Factor XIIIa Cross-Linking of the Marburg Fibrin: Formation of αm·γn-Heteromultimers and the α-Chain–Linked Albumin·γ Complex, and Disturbed Protofibril Assembly Resulting in Acquisition of Plasmin Resistance Relevant to Thrombophila

Blood ◽  
1998 ◽  
Vol 91 (9) ◽  
pp. 3282-3288 ◽  
Author(s):  
Teruko Sugo ◽  
Chizuko Nakamikawa ◽  
Mikihiro Takebe ◽  
Isao Kohno ◽  
Rudorf Egbring ◽  
...  
Keyword(s):  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Caesarian Section ◽  
Tissue Type ◽  
Factor Xiiia ◽  
Cross Linking ◽  
Amino Acid Residues ◽  
Pelvic Vein ◽  
Vein Thrombosis ◽  
Type Plasminogen Activator

Abstract The truncated Aα-chain of fibrinogen Marburg is partly linked with albumin by a disulfide bond. Based on the recovery of the first six amino acid residues assigned to the subunit polypeptides of fibrinogen (the Aα-and γ-chains) and albumin, 0.33 mol of albumin was estimated to be linked to 1 mol of the Marburg fibrinogen. When the Marburg fibrinogen was clotted with thrombin-factor XIIIa-Ca2+, various αmγnheteromultimers were produced, and part of the albumin was cross-linked to the γ-chain. Acid-solubilized Marburg fibrin monomer failed to form large aggregates that could be detected by monitoring turbidity at A350, but it was able to enhance tissue-type plasminogen-activator–catalyzed plasmin generation, though not as avidly as the normal control, indicating that the double-stranded protofibrils had, to some extent, been constructed. This idea seems to be supported by normal factor XIIIa–catalyzed cross-linking of the fibrin γ-chains. However, the cross-linked Marburg fibrin, being apparently fragile and translucent, was highly resistant against plasmin, and its subunit components were considerably retained for 48 hours as noted by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Although the exact mechanisms are still unclear, the albumin-incorporated factor XIIIa–cross-linked Marburg fibrin seems to have undergone a critical structural alteration(s) to acquire resistance against plasmin. This aquisition of plasmin resistance may be contributed to the postoperative pelvic vein thrombosis and recurrent pulmonary embolisms in the patient after caesarian section for her first delivery at the age of 20 years.

Localization of the α-Chain Cross-Linking Sites in Human Fibrinogen

1979 ◽  
Author(s):  
B. Hessel ◽  
G. Savidge ◽  
B. Blombäck
Keyword(s):  
Experimental Data ◽  
Calcium Ions ◽  
Whole Blood ◽  
Cyanogen Bromide ◽  
Degradation Products ◽  
Factor Xiiia ◽  
Antigenic Determinants ◽  
Cross Linking ◽  
Human Fibrinogen ◽  
The Cross

Antisera were raised in rabbits against the fragments Hi2-DSK and N-DSK obtained from cyanogen bromide treatment of human fibrinogen. Sensitive radioimmunoassay techniques were developed to quantitate substances sharing the same antigenic determinants as these fragments. We have found that the cross-linking process in fibrin formation involves the Hi2-DSK region which has its origin in the Aα-chain of fibrinogen. This relationship is based upon the following experimental data: - Fibrin samples produced in the presence and absence of Factor Xllla and Ca++ and thus cross-Linked and non cross-linked respectively were treated with cyanogen bromide. The resulting degradation products were chroraatographed on Sephadex G-100, and radioimmunoassays for Hi2-DSK and N-DSK were performed on the eluates. The elution profiles of Hi2-DSK and N-DSK antigenic determinants demonstrated that Hi2-DSK eluated after N-DSK in non cross-linked fibrin samples but in the cross-linked samples Hi2-DSK eluated before N-DSK. These results demonstrate that the Hi2-DSK region is involved in the cross-linking process in the presence of Factor XIIIa and calcium ions. Fibrin samples produced in the coagulation of whole blood showed a similar cross-linking profile.Supported by The Bank of Sweden Tercentenary Foundation.

Aggregated, Conformationally Changed Fibrinogen Exposes the Stimulating Sites for t-PA-Catalysed Plasminogen Activation

1996 ◽  
Vol 75 (02) ◽  
pp. 326-331 ◽  
Author(s):  
Unni Haddeland ◽  
Knut Sletten ◽  
Anne Bennick ◽  
Willem Nieuwenhuizen ◽  
Frank Brosstad
Keyword(s):  
Conformational Changes ◽  
Gel Permeation Chromatography ◽  
Tissue Type ◽  
Plasminogen Activation ◽  
Chromogenic Substrate ◽  
Type Plasminogen Activator ◽  
Gel Permeation ◽  
Self Association ◽  
Fibrin Monomers ◽  
Stimulation Of

SummaryThe present paper shows that conformationally changed fibrinogen can expose the sites Aα-(148-160) and γ-(312-324) involved in stimulation of the tissue-type plasminogen activator (t-PA)-catalysed plasminogen activation. The exposure of the stimulating sites was determined by ELISA using mABs directed to these sites, and was shown to coincide with stimulation of t-PA-catalysed plasminogen activation as assessed in an assay using a chromogenic substrate for plasmin. Gel permeation chromatography of fibrinogen conformationally changed by heat (46.5° C for 25 min) demonstrated the presence of both aggregated and monomeric fibrinogen. The aggregated fibrinogen, but not the monomeric fibrinogen, had exposed the epitopes Aα-(148-160) and γ-(312-324) involved in t-PA-stimulation. Fibrinogen subjected to heat in the presence of 3 mM of the tetrapeptide GPRP neither aggregates nor exposes the rate-enhancing sites. Thus, aggregation and exposure of t-PA-stimulating sites in fibrinogen seem to be related phenomena, and it is tempting to believe that the exposure of stimulating sites is a consequence of the conformational changes that occur during aggregation, or self-association. Fibrin monomers kept in a monomeric state by a final GPRP concentration of 3 mM do not expose the epitopes Aα-(148-160) and γ-(312-324) involved in t-PA-stimulation, whereas dilution of GPRP to a concentration that is no longer anti-polymerizing, results in exposure of these sites. Consequently, the exposure of t-PA-stimulating sites in fibrin as well is due to the conformational changes that occur during selfassociation.

Tridegin, a Novel Peptidic Inhibitor of Factor XIIIa from the Leech, Haementeria ghilianii, Enhances Fibrinolysis In Vitro

1997 ◽  
Vol 77 (05) ◽  
pp. 0959-0963 ◽  
Author(s):  
Lisa Seale ◽  
Sarah Finney ◽  
Roy T Sawyer ◽  
Robert B Wallis
Keyword(s):  
Potent Inhibitor ◽  
Platelet Rich Plasma ◽  
Factor Xiiia ◽  
Cross Linking ◽  
Fibrinolytic Enzymes ◽  
Small Polypeptide ◽  
Tissue Plasminogen ◽  
Protein Cross Linking

SummaryTridegin is a potent inhibitor of factor Xllla from the leech, Haementeria ghilianii, which inhibits protein cross-linking. It modifies plasmin-mediated fibrin degradation as shown by the absence of D-dimer and approximately halves the time for fibrinolysis. Plasma clots formed in the presence of Tridegin lyse more rapidly when either streptokinase, tissue plasminogen activator or hementin is added 2 h after clot formation. The effect of Tridegin is markedly increased if clots are formed from platelet-rich plasma. Platelet-rich plasma clots are lysed much more slowly by the fibrinolytic enzymes used and if Tridegin is present, the rate of lysis returns almost to that of platelet- free clots. These studies indicate the important role of platelets in conferring resistance to commonly used fibrinolytic enzymes and suggest that protein cross-linking is an important step in this effect. Moreover they indicate that Tridegin, a small polypeptide, may have potential as an adjunct to thrombolytic therapy.

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