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PubMed1978

An equilibrium study of metal ion binding to human plasma coagulation factor XIII.

1. The binding of Ca2+ to plasma coagulation Factor XIII from man and from cow caused a small decrease in the intrinsic fluorescence of the protein with a dissociation constant of 0.1 mM. A similar decrease was observed with the thrombin-activated Factors (Factors XIIa). The decrease in protein fluorescence was also caused by both Ni2+ and Mn2+ but not by Mg2+. 2. 45Ca2+ binding was directly demonstrated by equilibrium dialysis. Ca2+ at 0.2 mM bound to Factor XIII (a2b2) and Factor XIIIa (a'2b2) but not to isolated b2-protein. A tight-binding site for Ca2+ is associated with the a-subunits. 3. The Ca2+ essential for the enzyme activity of Factor XIII from man, pig and cow can be replaced by Ni2+, Cu2+, La3+, Mn2+, Fe3+, Y3+, Co2+, Sr2+ or Tb3+, but not by Mg2+.

Animals↗
PubMed1996

Changes in coagulation and fibrinolytic parameters caused by extracorporeal circulation.

During cardiopulmonary bypass (CPB) mechanical stress and the contact of blood with artificial surfaces lead to the activation of pro- and anticoagulant systems and the complement cascade, and to changes in cellular components. This phenomenon causes the "postperfusion-syndrome", with leukocytosis, increased capillary permeability, accumulation of interstitial fluid, and organ dysfunction. In this study, we focused on the influence of the extracorporeal circulation, sternotomy, and heparin administration on the activation of coagulation and fibrinolysis. In 15 patients we investigated coagulation parameters before, during and post CPB, i.e., fibrinogen, antithrombin (AT) III, thrombin-antithrombin complex (TAT), prothrombin fragments F1 + 2 (F1 + 2), factor (F) XIIa, tissue factor (TF), and parameters of the fibrinolytic system, i.e., plasmin-antiplasmin-complex (PAP), D-dimer, tissue-plasminogen-activator (tPA), urokinase-type plasminogen activator (uPA), and plasminogen-activator inhibitor type 1 (PAI 1). The results demonstrate distinct alterations in the above mentioned parameters. Despite administration of a high dose of heparin (activated clotting time [ACT] > 450s) combined with a low dose of aprotinin, activation of the coagulation and fibrinolytic pathways was observed. We found this activation was mainly caused by CPB and not by sternotomy. The activation of coagulation was due to foreign surface contact (F XII => F XIIa) as well as to an effect of tissue factor release in the late phase of CPB. The enhanced fibrinolytic activity during CPB was, at least in part, caused by tPA and was followed by PAI 1 release.

Anticoagulants↗
PubMed1987

Acetylcysteine in rats: inhibition of activation of prekallikrein and factor XII--protection against dextran-induced blood pressure fall.

Acetylcysteine (AC) injected intravenously into rats (200 mg/kg) had no effect on blood pressure, but significantly inhibited dextran-induced (40 mg/kg) blood pressure fall. Injection of AC also reversibly blocked the activation of prekallikrein (PK) normally obtained in plasma incubated with acetone. Kallikrein was assayed as plasminogen activator, S-2302 amidase and BAEe esterase. Also the activation of factor XII to factor XIIa, assayed as prekallikrein activator, was strongly inhibited in AC-treated rats. It is suggested that the partial blockade of dextran-induced shock is correlated with an inhibition of activation of PK and factor XII. Previous experiments had demonstrated an extensive, but reversible in vitro inhibition of human plasma kallikrein by AC. In view of such data it is concluded that the present results obtained with AC in rats are probably due to an inhibition of plasma kallikrein and its activation of factor XII.

Acetylcysteine↗
PubMed1988

Coagulation augments neutrophil C3b receptors via formation of a protein(s) unrelated to fibrinolysis or C5 activation.

The present study investigated the effect of coagulation on neutrophil complement receptors (CRs) 1 and 3, which are specific for the opsonins C3b and C3bi. Incubation of neutrophils in autologous serum, but not in plasma, increased the mean (+/- SD) expression of CR1 (x3.43 +/- 0.93) and CR3 (x3.07 +/- 0.86), in comparison with incubation in buffer. Serum also increased neutrophil superoxide production in response to opsonized zymosan from 0.48 +/- 0.21 to 1.05 +/- 0.25 nmol/10(6) cells/min. Similarly, calcium conversion of platelet-rich plasma (but not platelet-poor plasma) to serum also increased both CR1 and CR3 expression. This finding, as well as the fact that freeze-thawed platelet-rich plasma (but not platelet-poor plasma) increased CR expression, indicated that platelet constituents were the origin of this CR-inducing activity. Other nonplatelet factors formed during coagulation, such as C5a, fibrinogen degradation products, kallikrein, and factor XIIa, were shown not to be responsible for this CR-inducing activity.

Blood Coagulation Factors↗
PubMed1995

Plasmin-mediated activation of contact system in response to pharmacological thrombolysis.

BACKGROUND: Thrombin activity increases in patients treated with coronary thrombolysis for acute myocardial infarction, but the mechanisms are not well defined. We have shown that thrombin activity increases in plasma and whole blood incubated with plasminogen activators and appears to be plasmin mediated and dependent on activity of the factor VIIIa/IXa complex. METHODS AND RESULTS: In the present study, increases in thrombin activity induced by incubation of recalcified citrated plasma with 0.16 to 0.5 mumol/L plasmin at 37 degrees C were markedly attenuated in recalcified citrated plasma deficient in factors XI or XII, prekallikrein, or high molecular weight kininogen, as well as in plasma incubated with plasmin in the presence of 3.5 mumol/L corn trypsin inhibitor, a specific factor XIIa inhibitor. Increases in thrombin activity also occurred in nonanticoagulated whole blood incubated with pharmacological concentrations of plasminogen activators and were markedly attenuated in the presence of corn trypsin inhibitor. Plasmin-mediated (0.25 mumol/L) activation of purified factor XII occurred in 0.05 mol/L Tris-HCl and 0.012 mol/L NaCl (pH 7.8) at 37 degrees C, resulting in equimolar quantities of two fragments that corresponded to cleavage of factor XII at Arg353-Val354, the site involved in kallikrein-mediated activation of factor XII, and cleavage at Lys346-Ser347, an apparently novel site of plasmin-mediated hydrolysis of factor XII. Contact activation was also demonstrated in plasma samples from patients after treatment with fibrinolytic agents for myocardial infarction, by demonstrating cleavage of high molecular weight kininogen from its one-chain to its two-chain form by ligand blotting with 125I-prekallikrein. CONCLUSIONS: Plasmin-mediated activation of the contact system of coagulation appears to account, at least in part, for increases in procoagulant activity in patients treated with fibrinolytic agents. It may also explain hypotension, by release of bradykinin from high molecular weight kininogen, and complement activation, by activated factor XII, that has been demonstrated in these patients.

Biomarkers↗
PubMed2004

Accelerating ability of synthetic oligosaccharides on antithrombin inhibition of proteinases of the clotting and fibrinolytic systems. Comparison with heparin and low-molecular-weight heparin.

The abilities of three synthetic oligosaccharides to accelerate antithrombin inhibition of ten clotting or fibrinolytic proteinases were compared with those of unfractionated, fractionated high-affinity and low-molecular-weight heparins. The results show that the anticoagulant effects of the latter three heparins under conditions approximating physiologic are exerted almost exclusively by acceleration of the inactivation of thrombin, factor Xa and factor IXa to near diffusion-controlled rate constants of approximately 10(6) - 10(7) M(-1).s(-1). All other proteinases are inhibited with at least 20-fold lower rate constants. The anti-coagulant ability of the synthetic regular (fondaparinux) and high-affinity (idraparinux) pentasaccharides is due to a common mechanism, involving acceleration of only factor Xa inhibition to rate constants of approximately 10(6) M(-1).s(-1) . A synthetic hexadecasaccharide, containing both the pentasaccharide sequence and a proteinase binding site, exerts its anticoagulant effect by accelerating antithrombin inactivation of both thrombin and factor Xa to rate constants of approximately 10(6) - 10(7) M(-1).s(-1), although thrombin appears to be the more important target. In contrast, factor IXa inhibition is appreciably less stimulated. The conformational change of antithrombin induced both by the pentasaccharides and longer heparins contributes substantially, approximately 150-500-fold, to accelerating the inactivation of factors Xa, IXa and VIIa and moderately, approximately 50-fold, to that of factor XIIa and tissue plasminogen activator inhibition. The bridging effect due to binding of antithrombin and proteinase to the same, long heparin chain is dominating, approximately 1000-3000-fold, for thrombin inhibition and is appreciably smaller, although up to approximately 250-350-fold, for the inactivation of factors IXa and XIa. These results establish the proteinase targets of heparin derivatives currently used in or considered for thrombosis therapy and give new insights into the mechanism of heparin acceleration of antithrombin inhibition of proteinases.

Antithrombin III↗
PubMed2004

Factor VIIa-mediated tenase function on activated platelets under flow.

BACKGROUND: Tissue factor (TF) and/or active factor (F)VIIa may be stored inside resting platelets. OBJECTIVES: The objective of this study was to examine if platelets, following activation of GPVI, could support tenase and prothrombinase activity without any exogenously added tissue factor. METHODS: Thrombin (IIa) formation on gel-filtered platelets with added factors or the clotting of platelet-free plasma (PFP) or platelet-rich plasma (PRP) supplemented with corn trypsin inhibitor (CTI) (to inhibit factor XIIa) was studied in well plate assays with a fluorogenic thrombin substrate or in flow assays by fibrin visualization. RESULTS: Pretreatment of convulxin (CVX)-stimulated, fibrinogen-adherent, gel-filtered platelets with anti-TF, anti-FVII/VIIa, or 1 nm PPACK [inhibitor of FVIIa, factor XIa and factor (F)IIa] delayed fibrin deposition on platelets perfused with PFP/CTI at 62.5 s(-1). Anti-TF or anti-FVII/VIIa also attenuated thrombin generation in plate assays using recalcified PRP/CTI treated with CVX. Anti-TF or anti-FVII/VIIa (but not inhibited factor IXa) delayed the burst in thrombin production by gel-filtered platelets suspended in prothrombin and CVX by 14 min and 40 min, respectively. Anti-FVII/VIIa completely eliminated thrombin generation on fibrinogen-adherent, gel-filtered platelets pretreated with 10 micro m PPACK and 10 micro m EGR-CK [inhibitor of factor (F)Xa], rinsed, and then supplemented with CVX, prothrombin, and FX. Addition of anionic phospholipid to PFP/CTI or to a mixture of prothrombin, FX, and recVIIa was not sufficient to generate detectable tenase activity. Lastly, isolated, unactivated neutrophils suspended in FX, FII and recVIIa supported a very low level of thrombin generation sensitive to antagonism of P-selectin, CD18, and TF. CONCLUSIONS: Activated platelets supported tenase and prothrombinase activity by elevating the function or level of FVIIa and exposing active FVIIa or FVIIa-cofactor(s), distinct from anionic lipid, that may be, in part, TF.

Animals↗
PubMed1986

Amino acid sequence of human factor XI, a blood coagulation factor with four tandem repeats that are highly homologous with plasma prekallikrein.

A lambda gtll cDNA library prepared from human liver poly(A) RNA has been screened with affinity-purified antibody to human factor XI, a blood coagulation factor composed of two identical polypeptide chains linked by a disulfide bond(s). A cDNA insert coding for factor XI was isolated and shown to contain 2097 nucleotides, including 54 nucleotides coding for a leader peptide of 18 amino acids and 1821 nucleotides coding for 607 amino acids that are present in each of the 2 chains of the mature protein. The cDNA for factor XI also contained a stop codon (TGA), a potential polyadenylation or processing sequence (AACAAA), and a poly(A) tail at the 3' end. Five potential N-glycosylation sites were found in each of the two chains of factor XI. The cleavage site for the activation of factor XI by factor XIIa was identified as an internal peptide bond between Arg-369 and Ile-370 in each polypeptide chain. This was based upon the amino acid sequence predicted by the cDNA and the amino acid sequence previously reported for the amino-terminal portion of the light chain of factor XI. Each heavy chain of factor XIa (369 amino acids) was found to contain 4 tandem repeats of 90 (or 91) amino acids plus a short connecting peptide. Each repeat probably forms a separate domain containing three internal disulfide bonds. The light chains of factor XIa (each 238 amino acids) contain the catalytic portion of the enzyme with sequences that are typical of the trypsin family of serine proteases. The amino acid sequence of factor XI shows 58% identity with human plasma prekallikrein.

Amino Acid Sequence↗
PubMed1992

Additive and synergistic pharmacologic inhibition of equine fibrinoligase (factor XIIIa*-like) biochemical activity.

A selected group of pharmaceutical compounds were evaluated for the ability to inhibit the biochemical activity of fibrinoligase (coagulation factor XIIIa*) in pooled equine plasma. Criteria for the pharmaceuticals selected were based on the mechanism of the transglutamination biochemical reaction mediated by coagulation factor XIIa*. These criteria were complemented by recognition of the molecular configuration and chemical composition of amino acid residue side chains involved in the process of covalent fibrin monomer polymerization (cross-linking, transglutamination) mediated by this enzyme. Each pharmaceutical was evaluated individually and in combination with other potential coagulation factor XIIIa* inhibitors in an effort to detect additive and synergistic phenomenon. In this context, pharmaceuticals with a carbonylamide (eg, cefuroxime, Girard's reagent-P, prolinamide) were applied in concert with compounds with a terminal amine (eg, D-arginine, L-lysine) functional group. In concept, this method theoretically served to competitively simulate glutamine and lysine amino acid residues within strands of fibrin monomer substrate involved in phase I (carbonylamide) and phase II (terminal amine) of the transglutamination reaction (covalent fibrin monomer cross-linking). Halogen-dinitro and ethylene compounds were also evaluated because of their reported ability to inactivate enzyme systems dependent on an intact sulfhydryl group located at their biochemically active site (eg, cystine amino acid residue). This group of pharmaceutical compounds failed to inhibit the biochemical activity mediated by coagulation factor XIIIa* in equine plasma.

Animals↗
PubMed1995

Monoclonal antibody F1 binds to the kringle domain of factor XII and induces enhanced susceptibility for cleavage by kallikrein.

In a previous study we have shown that monoclonal antibody F1 (MoAb F1), directed against an epitope on the heavy chain of factor XII distinct from the binding site for anionic surfaces, is able to activate factor XII in plasma (Nuijens JH, et al: J Biol Chem 264; 12941, 1989). Here, we studied in detail the mechanism underlying the activation of factor XII by MoAb F1 using purified proteins. Formation of factor XIIa was assessed by measuring its amidolytic activity towards the chromogenic substrate H-D-Pro-Phe-Arg-pNA (S-2302) in the presence of soybean trypsin inhibitor and by assessing cleavage on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Upon incubation with MoAb F1 alone, factor XII was auto-activated in a time-dependent fashion, activation being maximal after 30 hours. Factor XII incubated in the absence of MoAb F1 was hardly activated by kallikrein, whereas in the presence of MoAb F1, but not in that of a control MoAb, the rate of factor XII activation by kallikrein was promoted at least 60-fold. Maximal activation of factor XII with kallikrein in the presence of MoAb F1 was reached within 1 hour. This effect of kallikrein on the cleavage of factor XII bound to MoAb F1 was specific because the fibrinolytic enzymes plasmin, urokinase, and tissue-type plasminogen activator could not substitute for kallikrein. Also, trypsin could easily activate factor XII, but in contrast to kallikrein, this activation was independent of MoAb F1. SDS-PAGE analysis showed that the appearance of amidolytic activity correlated well with cleavage of factor XII. MoAb F1-induced activation of factor XII in this purified system was not dependent on the presence of high-molecular-weight kininogen (HK), in contrast to the activation of the contact system in plasma by MoAb F1. Experiments with deletion mutants revealed that the epitopic region for MoAb F1 on factor XII is located on the kringle domain. Thus, this study shows that binding of ligands to the kringle domain, which does not contribute to the proposed binding site for negatively charged surfaces, may induce activation of factor XII. Therefore, these findings point to the existence of multiple mechanisms of activation of factor XII.

Amino Acid Sequence↗
PubMed1997

Activation of coagulation and platelets is affected by the hydrophobicity of artificial surfaces.

Clot formation is a limiting factor in the use of biomaterials. We investigated the effect of surface hydrophobicity on haemostatic activation in vitro, using five polyetherurethanes of varying surface hydrophobicity (C94, C74, C54 and C34), C94 the most, and C34, the least hydrophobic, and compared them with a commercial standard pellethane. Sterilised sacks were filled with heparinised blood, rotated at 37 degrees C for 24 h and sequential samples collected into 0.103 M sodium citrate. Thrombin generation measured by thrombin-antithrombin III complexes showed a difference between the polymers at 3 h through to 6 h (P < 0.05), C94 showing the least activation and C34 the most. Factor XIIa and D-dimer levels increased between 12 (P < 0.05) and 24 h (P < 0.01) for all polyetherurethanes. The ratio of soft:hard segments (which determine hydrophobicity) of the polyetherurethanes showed a direct relationship with the degree of activation of coagulation and fibrinolysis. There was no significant increase in monocyte tissue factor expression at 5 and 105 min. Platelet function as measured by whole blood platelet aggregation showed a reduction with pellethane and C94 after 1 h using collagen, with no changes for C34, Altering surface hydrophobicity has diverse effects on haemostatic pathways, with the most hydrophobic surfaces causing least activation of coagulation but most activation of platelets.

Adult↗
PubMed1989

Factor XIIIa-mediated cross-linking of fibronectin in fibroblast cell layers. Cross-linking of cellular and plasma fibronectin and of amino-terminal fibronectin fragments.

Factor XIIIa cross-links plasma fibronectin as it is being assembled into the extracellular matrix of cultured human skin fibroblasts (Barry, E. L. R., and Mosher, D. F. (1988) J. Biol. Chem. 262, 10464-10469). We have further characterized this process. Fibroblasts were metabolically labeled with proline in the presence or absence of ascorbate and Factor XIIIa. Endogenous fibronectin in the extracellular matrix was cross-linked by Factor XIIIa. There was no evidence for cross-linking of collagenous proteins. Fibro-blast cell layers were incubated with iodinated 27-kDa heparin-binding or 70-kDa collagen- and heparin-binding amino-terminal fibronectin fragments. Factor XIIa cross-linked the fragments into high molecular weight aggregates. The amounts of cross-linked fragments reaches a steady state after 1 to 2 h, whereas intact fibronectin continues to be cross-linked for 24 h. When fibroblast cell layers were pulsed with iodinated fibronectin or amino-terminal fragments and Factor XIIIa was included in the chase media, the high molecular weight aggregates were formed in a step-wise manner. The smallest cross-linking steps were to high molecular weight extracellular matrix molecules forming approximately 270-, 300-, and 440-kDa complexes for the 27-kDa fragment, 70-kDa fragment, and intact fibronectin, respectively. When iodinated fibronectin was bound to fibroblast cell layers and chased into the matrix pool in the absence of Factor XIIIa, it could also be cross-linked into high molecular weight complexes when Factor XIIIa was added to the media. These results, therefore, indicate that both cellular and plasma fibronectin and amino-terminal fragments are cross-linked specifically by Factor XIIIa, that the cross-linking is probably to other fibronectin molecules rather than to collagenous proteins, and that both assembling and assembled fibronectin are substrates for Factor XIIIa.

Collagen↗
PubMed1984

Effects of cryoglobulin on fibrin clot formation and fibrinolysis.

It was confirmed that activation of the kallikrein-kinin enzyme system in cryoglobulinemia might be initiated by activation of factor XII to factor XIIa by cryoglobulin. It was also demonstrated that cryoglobulin or fibrin clots lost their ability to redissolve on warming to 37 degrees C, and consequently the lysis time of fibrin clots was increased.

Blood Coagulation↗
PubMed2002

Contact system. New concepts on activation mechanisms and bioregulatory functions.

This review considers the data of recent years concerning the contact system initiating the activation of blood plasma proteolytic systems, such as hemocoagulation, fibrinolysis, kininogenesis, and also complement and angiotensinogenesis. The main proteins of the contact system are the factors XII and XI, prekallikrein, and high-molecular-weight kininogen. The data on the structure, functions, and biosynthesis of these proteins and on their genes are presented. Studies in detail on the protein-protein interactions during formation of the ensemble of the contact system components on the anionic surface resulted in the postulation of the mechanism of activation of this system associated with generation of the XIIa factor and of kallikrein. This mechanism is traditionally considered a trigger of processes for the internal pathway of the hemocoagulating cascade. However, the absence of direct confirmation of such activation in vivo and the absence of hemorrhagia in the deficiency of these components stimulated the studies designed to find another mechanism of their activation and physiological role outside of the hemostasis system. As a result, a new concept on the contact system activation on the endothelial cell membrane was proposed. This concept is based on the isolation of a complex of proteins, which in addition to the above-mentioned proteins includes cytokeratin 1 and the receptors of the urokinase-like plasminogen activator and of the complement q-component. The ideas on the role of this system in the biology of vessels are developed. Some of our findings on the effect of leukocytic elastase on the key components of the contact system are also presented.

Blood↗
PubMed1975

On the activation of bovine plasma factor XIII. Amino acid sequence of the peptide released by thrombin and the terminal residues of the subunit polypeptides.

A blood coagulation factor, Factor XIII, was highly purified from bovine fresh plasma by a method similar to those used for human plasma Factor XIII. The isolated Factor XIII consisted of two subunit polypeptides, a and b chains, with molecular weights of 79,000 +/- 2,000 and 75,000 +/- 2,000, respectively. In the conversion of Factor XIII to the active enzyme, Factor XIIIa, by bovine thrombin [EC 3.4.21.5], a peptide was liberated. This peptide, designated tentatively as "activation peptide," was isolated by gel-filtration on a Sephadex G-75 column. It contained a total of 37 amino acid residues with a masked N-terminal residue and C-terminal arginine. The whole amino acid sequence of "Activation peptide" was established by the dansyl-Edman method and standard enzymatic techniques, and the masked N-terminal residue was identified as N-acetylserine by using a rat liver acylamino acid-releasing enzyme. This enzyme specifically cleaved the N-acetylserylglutamyl peptide bond serine and the remaining peptide, which was now reactive to 1-dimethylamino-naphthalene-5-sulfonyl chloride. A comparison of the sequences of human and bovine "Activation peptide" revealed five amino acids replacements, Ser-3 to Thr; Gly-5 to Arg; Ile-14 to Val; Thr-18 to Asn, and Pro-26 to Leu. Another difference was the deletion of Leu-34 in the human peptide. Adsorption chromatography on a hydroxylapatite column in the presence of 0.1% sodium dodecyl sulfate was developed as a preparative procedure for the resolution of the two subunit polypeptides, a or a' chain and b chain, constituting the protein molecule of Factor XIII or Factor XIIIa. End group analyses on the isolated pure chains revealed that the structural change of Factor XIII during activation with thrombin occurs only in the N-terminal portion of the a chain, not in the N-terminal end of the b chain or in the C-terminal ends of the a and b chains. From these results, it was concluded that the activation of bovine plasma Factor XIII by thrombin must be accompanied by a limited proteolysis of the arginyl-glycyl bond located in the N-terminal region of the a chain, liberating the "Activation peptide." The possibility of activating Factor XII with other porteinases was examined using Factor Xa [EC 3.4.21.6], Factor XIIa, kallikreins [EC 3.4.21.8], urokinase [EC 3.4.99.26], trypsin [EC 3.4.21.4], ficin [EC 3.4.22.3], papain [EC 3.4.22.2], and bromelain [EC 3.4.22.4]. Among these enzymes, only bromelain and trypsin showed clear activating effects.

Amino Acid Sequence↗
PubMed1986

Human plasma prekallikrein, a zymogen to a serine protease that contains four tandem repeats.

The amino acid sequence of human plasma prekallikrein was determined by a combination of automated Edman degradation and cDNA sequencing techniques. Human plasma prekallikrein was fragmented with cyanogen bromide, and 13 homogeneous peptides were isolated and sequenced. Cyanogen bromide peptides containing carbohydrate were further digested with trypsin, and the peptides containing carbohydrate were isolated and sequenced. Five asparagine-linked carbohydrate attachment sites were identified. The sequence determined by Edman degradation was aligned with the amino acid sequence predicted from cDNAs isolated from a lambda gt11 expression library. This library contained cDNA inserts prepared from human liver poly(A) RNA. Analysis of the cDNA indicated that human plasma prekallikrein is synthesized as a precursor with a signal peptide of 19 amino acids. The mature form of the protein that circulates in blood is a single-chain polypeptide of 619 amino acids. Plasma prekallikrein is converted to plasma kallikrein by factor XIIa by the cleavage of an internal Arg-Ile bond. Plasma kallikrein is composed of a heavy chain (371 amino acids) and a light chain (248 amino acids), and these 2 chains are held together by a disulfide bond. The heavy chain of plasma kallikrein originates from the amino-terminal end of the zymogen and is composed of 4 tandem repeats that are 90 or 91 amino acid residues in length. These repeat sequences are also homologous to those in human factor XI. The light chain of plasma kallikrein contains the catalytic portion of the enzyme and is homologous to the trypsin family of serine proteases.

Amino Acid Sequence↗
PubMed1989

Plasminogen activators in dextran sulfate-activated euglobulin fractions: a molecular analysis of factor XII- and prekallikrein-dependent fibrinolysis.

To elucidate the mechanism by which activation of the contact system of blood coagulation leads to expression of fibrinolytic activity, we have determined the molecular characteristics of the plasminogen activators present in dextran sulfate-treated euglobulin fractions by electrophoretic-zymographic analysis and specific immunoadsorption. In addition to free and protease inhibitor-bound tissue-type plasminogen activator (t-PA), dextran sulfate precipitates of euglobulins contained the complex formed between plasma kallikrein and C1-inhibitor, an indicator of prekallikrein activation. These precipitates also contained substantial fibrinolytic activity related to urinary-type plasminogen activator (u-PA). Autoradiographic analysis was then used to evaluate the cleavage of 125I-single-chain u-PA (prourokinase) in dextran sulfate euglobulins as well as after exposure to kallikrein or beta-factor XIIa. This analysis supported the conclusion that plasma kallikrein-mediated cleavage and activation of single-chain u-PA is the mechanism operative for the development of lytic activity in euglobulin precipitates following activation of the contact system.

Blood Proteins↗
PubMed1996

Nafamostat mesilate reduces blood-foreign surface reactions similar to biocompatible materials.

BACKGROUND: Nafamostat mesilate (FUT-175) is a synthetic serine protease inhibitor that inactivates coagulation, fibrinolysis, and platelet aggregation. Nafamostat mesilate may suppress the blood-foreign surface reaction similar to biocompatible materials by blocking factor XIIa. METHODS: We performed an in vitro study of cardiopulmonary bypass (CPB) with fresh human blood among the following three groups: standard CPB sets (C), biocompatible CPB sets (B), and standard CPB sets with FUT-175 (10 mg/L) (F). A clinical study using these same CPB groups also was performed in 45 patients undergoing aortocoronary bypass operations (15 patients each). We injected FUT-175 at 40 mg/h during CPB. RESULTS: In the in vitro study, both groups B and F showed significantly lower levels of coagulation factors, thrombin-antithrombin III complex, fibrinopeptide A, beta-thromboglobulin, complement C3a, granulocyte elastase, and free hemoglobin than group C at the conclusion of the study. Thrombin-antithrombin III complex and free hemoglobin in group F also were lower than in group B. The platelet count remained at a higher level in group F than in the other groups. Separation of bradykinin was suppressed most significantly in group F. In the clinical study, group F also showed significantly lower levels of alpha 2-plasmin inhibitor plasmin complex and C3a than both groups C and B. There were minimal levels of free hemoglobin in group F. CONCLUSIONS: Nafamostat mesilate may contribute major beneficial effects toward conservation of blood during CPB and prevention of coagulopathy after CPB.

Benzamidines↗