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:3.4.21.7 (plasmin)
9,023 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

This report describes a plasma prekallikrein assay which, unlike methods that employ contact activation, is not affected by the factor XII or HMW kininogen content of the plasma analyzed. In this assay beta-XIIa, a potent fluid-phase activator of prekallikrein, is added to diluted plasma in the presence of 20% acetone (to inactivate kallikrein inhibitors) at 30 degrees C and the kallikrein generated is measured with the chromogenic substrate S-2302. Prekallikrein is fully activated under these conditions and the activity remains stable for at least 30 hr. The mean prekallikrein concentration in plasma samples from 24 healthy individuals was 1.50 +/- 0.35 (S.D.) S-2302 U/ml, corresponding to 20.3 +/- 4.7 micrograms/ml prekallikrein (the specific activity of highly purified human prekallikrein was determined to be 74 S-2302 U/mg). In contrast, the mean concentration in five plasma samples from patients deficient in HMW kininogen was 0.38 +/- 0.02 S-2302 U/ml. No activity was generated in prekallikrein-deficient plasma, and essentially normal levels (1.35 +/- 0.18 S-2302 U/ml) were measured in plasmas from three patients with factor XII deficiency. Plasma prekallikrein was also quantitated by radial immunodiffusion, which gave results similar to those obtained by functional assay with beta-XIIa. The determination of plasma prekallikrein by direct activation with beta-XIIa in the presence of acetone offers several advantages over the use of contact activators such as dextran sulfate. These advantages include complete inactivation of kallikrein inhibitors and total activation of prekallikrein (even in plasmas deficient in other contact factors) without simultaneous generation of plasmin.
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PMID:Plasma prekallikrein: quantitative determination by direct activation with Hageman factor fragment (beta-XIIa). 654 73

We report the isolation of a specific protease zymogen from chicken plasma. The purification procedure involves barium citrate precipitation, ammonium sulfate fractionation, removal of plasminogen and plasmin on lysine-Sepharose, followed by anion and cation exchange, and gel permeation chromatography. Based on quantitative radioimmunoassay the zymogen is present in plasma at a concentration of 160 mg/liter, and it is obtained by our procedure in highly purified form with a yield of 1.4%. The single polypeptide chain contains an NH2-terminal alanine residue. The native molecule migrates in sodium dodecyl sulfate-polyacrylamide gel electrophoresis with an apparent molecular weight of 84,000 under reducing conditions. It can be identified as an inactive proenzyme because it has very low amidolytic activity, does not react with the fluorescent active site titrant 4-methyl-lumbelliferyl p-guanidinobenzoate, and does not incorporate radioactive [3H]diisopropylfluorophosphate. It is very susceptible to limited proteolysis which converts it to an active enzyme with trypsin-like specificity. The active enzyme, likewise a single polypeptide chain, migrates as a doublet with apparent molecular weights of 39,000 and 40,000. Its amidolytic activity with synthetic peptide substrates is at least 40-fold higher than that of the proenzyme, it reacts efficiently with 4-methylumbelliferyl p-guanidinobenzoate, and incorporates [3H]diisopropylfluorophosphate while undergoing irreversible inactivation. The enzyme appears to be a reasonably efficient plasminogen activator in zymographic gels, but not in solution. With human high molecular weight kininogen as substrate the enzyme was about 25% as efficient as human plasma kallikrein. It lacks any plasminogen-independent proteolytic activity with other protein substrates, and it hydrolyzes small peptide substrates designed for both human kallikrein and urinary urokinase, respectively. Inhibition studies with peptide chloromethyl ketones indicate enzymatic properties closer to human plasma kallikrein than to the human plasminogen activator urokinase (EC 3.4.21.31). The chicken plasma enzyme and the plasminogen activator from the conditioned media of Rous sarcoma virus-transformed chick embryo fibroblasts treated with tumor promoter are different by criteria of tryptic peptide maps, and amino acid composition and enzymatic specificity. The designations chicken plasma prekallikrein plasminogen proactivator and chicken plasma kallikrein plasminogen activator are proposed for the zymogen and enzyme forms, respectively. Using rabbit antibodies against the proenzyme we developed a solid phase immunoadsorption procedure that allowed us to isolate the protein with an overall yield of 11.4%.
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PMID:A proenzyme from chicken plasma similar to human plasma prekallikrein. 655 13

Activation of bovine plasma prekallikrein was investigated with several proteinases. Highly purified bovine plasma prekallikrein was rapidly activated to kallikrein [EC 3.4.21.8] by bovine activated Hageman factor, trypsin [EC 3.4.21.4] and Pronase P (proteinases from Streptomyces griseus) and more gradually by papain [EC 3.4.22.2] and ficin [EC 3.4.22.3]. Activation of prekallikrein was also observed with bovine plasmin [EC 3.4.21.7], but not with bovine clotting factors Xa (Stuart factor) [EC 3.4.21.6] and IXa (Christmas factor) or thrombin [EC 3.4.21.5]. Urokinase [EC 3.4.99.26], Reptilase, collagenase [EC 3.4.24.3], elastase [EC 3.4.21.11], alpha-chymotrypsin [EC 3.4.21.1], Nagarse [EC 3.4.21.14], and stem bromelain [EC 3.4.22 4] did not convert prekallikrein to kallikrein. Plasma kallikrein activated to Hageman factor released kinin rapidly from bovine high molecular weight (HMW) kininogen. However, from bovine low molecular weight (LMW) kininogen, liberation of kinin was extremely slow. The kallikrein activity was inhibited by soybean trypsin inhibitor (SBTI), Trasylol, diisopropylfluorophosphate (DFP), and N-alpha-tosyl-L-lysine chloromethylketone (TLCK), but not by egg-white trypsin inhibitor (EWTI), lima bean trypsin inhibitor (LBTI), heparin or hexadimethrine bromide (Polybrene). The kallikrein formed an enzyme-inhibitor complex with SBTI and Trasylol, but not with LBTI. Prekallikrein did not react with SBTI. Prekallikrein consists of a single polypeptide chain of molecular weight about 90,000, as estimated by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. Activation of prekallikrein by Hageman factor was found to involve cleavage of the single peptide bond on the disulfide-bridged polypeptide chain, and no change of molecular weight was observed during the activation. The peptide bond cleaved in prekallikrein by the activation was an Arg-X peptide bond on a disulfide-bridged polypeptide chain.
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PMID:Studies on prekallikrein of bovine plasma. II. Activation of prekallikrein with proteinases and properties of kallikrein activated by bovine Hageman factor. 676 24

Incubation of normal human plasma or its euglobulin fraction with chloroform enhances the generation of proteolytic activity attributable to plasmin. The mechanisms involved remain unclear. Earlier, plasmin formation was found to be impaired in the chloroform-treated euglobulin fraction of Hageman trait plasma. We now demonstrate sharply reduced generation of fibrinolytic and caseinolytic activities in prekallikrein-deficient (Fletcher trait) and HMW kininogen-deficient (Fitzgerald trait) plasmas as well. These defects were corrected by addition of the missing factors. In addition, precipitation of the euglobulin fraction of plasma activated small but measurable amounts of HF. With the use of 125I-HF, cleavage of the native molecule into its amino-terminal and carboxy-terminal fragments occurred pari passu with the generation of plasmin. These experiments suggest that one pathway through which HF can bring about formation of plasmin in chloroform-treated euglobulin fractions involves the participation of plasma prekallikrein and HMW kininogen, presumably through mechanisms similar to those evoked by surface activation of HF.
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PMID:Chloroform-induced fibrinolysis. Its dependence on Hageman factor, plasma prekallikrein, and high molecular weight kininogen. 735 11

Activation of the fibrinolytic, coagulation and plasma kallikrein-kinin systems may be responsible for some of the coagulation disorders and inflammatory sequelae seen after extracorporeal circulation. The activation pattern of these systems was studied in 10 children undergoing open heart surgery with extracorporeal circulation. Blood samples were drawn serially before, during and up to 48 h after surgery. The heparin injection induced a significant elevation of plasmin (PL) (p < 0.05) which stayed elevated during extracorporeal circulation. Antiplasmin (AP) values were reduced at wound closure, while the levels were significantly elevated 48 h postoperatively (p < 0.05). alpha 2-antiplasmin-plasmin (APP) increased significantly perioperatively peaking 10 min after the initiation of cardiopulmonary bypass (p < 0.05). The coagulation markers thrombin-antithrombin (TAT) and the prothrombin fragment F1 & 2 increased significantly, peaking at wound closure and at termination of bypass respectively (p < 0.05). Plasma kallikrein (KK) values increased significantly with subsequent decreased levels of prekallikrein (PKK) and kallikrein inhibitor (KKI) after heparin injection. The KK level stayed elevated during cardiopulmonary bypass (CPB). The proenzyme functional inhibition index (PFI index), defined as the sum of deviations from the control values for proenzyme and functional inhibition values of the coagulation, fibrinolytic and plasma kallikrein-kinin systems, correlated significantly to the duration of cardiopulmonary bypass (p < 0.05). We conclude that open heart surgery in children activates the fibrinolytic, coagulation and plasma kallikrein-kinin systems.
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PMID:Activation of the fibrinolytic, coagulation and plasma kallikrein-kinin systems during and after open heart surgery in children. 756 39

Clots formed from platelet rich plasma were found to be lysed more readily by low concentrations of pro-urokinase (pro-UK) than clots formed from platelet poor plasma. This was not a non-specific effect since the reverse occurred with tissue plasminogen activator. A mechanical explanation due to platelet-mediated clot retraction was excluded by experiments in which retraction was inhibited with cytochalasin B. Therefore, a platelet-mediated enzymatic mechanism was postulated to explain the promotion of fibrinolysis. Casein autography of isolated platelets revealed a approximately 90 kDa band of activity which comigrated with plasma prekallikrein (PK)/kallikrein, a known activator of pro-UK. Furthermore, treatment of platelets with plasma PK activator (PPA), consisting essentially of factor XIIa, induced activation of pro-UK and of chromogenic substrate for kallikrein (S-2302). This activity corresponded to approximately 40-200 pM kallikrein per 10(8) washed and gel filtered platelets per ml. The activation of pro-UK by PPA-pretreated platelets was dose-dependent and inhibited by soybean trypsin inhibitor but not by bdellin, a specific inhibitor of plasmin, nor by the corn inhibitor of factor XIIa. Kinetic analysis of pro-UK activation by kallikrein showed promotion of the reaction by platelets. The KM of the reaction was reduced by platelets by approximately 7-fold, while the kcat was essentially unchanged. In conclusion, PK was shown to be tightly associated with platelets where it can be activated by factor XIIa during clotting. The activation of pro-UK by platelet-bound kallikrein provides an explanation for the observed platelet mediated promotion of pro-UK-induced clot lysis.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Platelet-bound prekallikrein promotes pro-urokinase-induced clot lysis: a mechanism for targeting the factor XII dependent intrinsic pathway of fibrinolysis. 802

Type IV collagenases are secreted as latent 92 and 72 kDa proenzymes which are then activated extracellularly. The mechanisms by which they are activated in vivo are not clear. We have studied the activation of porcine endothelial cell type IV collagenases by tissue and plasma kallikrein, and found that tissue kallikrein was a very efficient activator of the 92 kDa type IV collagenase. Enzyme cleavage was observed at concentrations of tissue kallikrein as low as 0.1 microgram/ml. Plasma kallikrein had no effect. By comparison, plasmin, which has been proposed to be the physiological activator of interstitial collagenase and stromelysin, and elastase were much less effective, and high concentrations (plasmin at 100-200 micrograms/ml and elastase at 20 micrograms/ml) were required to cause only a limited cleavage which was not associated with an increase in activity, as observed by the gelatin-gel lysis assay. In addition tissue kallikrein was found by immunohistochemistry to be present in the extracellular matrix of the intima of porcine aortic vessel wall. These findings suggest that tissue kallikrein can be a potential activator of the 92 kDa type IV collagenase in vivo.
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PMID:Activation of the 92 kDa type IV collagenase by tissue kallikrein. 825 70

The aim of this study was to evaluate status of plasma kinin system in patients with ovarian hyperstimulation syndrome (OHSS), in order to investigate whether activation of the plasma kinin system correlates with increased blood coagulability. In the first part of the study, concentrations of plasma prekallikrein (PK) in OHSS cycles (n = 13) were monitored from the day of human chorionic gonadotrophin (HCG) administration to the mid-luteal phase, and were compared with those of control cycles (n = 17). The average value of PK in OHSS cycles began to decrease on day 8, and by day 10 was significantly lower than that of control cycles (86 +/- 6 versus 106 +/- 4%, P <0.01). The time course of changes in PK concentration correlated well with the clinical condition of OHSS patients. In the second part of the study, we obtained data from 26 patients who were hospitalized because of severe OHSS, to investigate the correlation between PK and other haemostatic markers. OHSS patients with severe PK reduction (<80% normal, n = 9) demonstrated significantly higher values of plasma thrombin-antithrombin III and plasmin-alpha2 antiplasmin, and more severe haemoconcentration, compared to those OHSS patients who had no reduction in PK (n = 17). In conclusion, our data suggest that activation of the plasma kinin system occurs specifically and occasionally in OHSS patients, and is associated with increased blood coagulability. Thus, when an OHSS patient demonstrates a low value of plasma PK, more careful management is required to prevent thromboembolic complications.
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PMID:Activation of plasma kinin system correlates with severe coagulation disorders in patients with ovarian hyperstimulation syndrome. 919 35

A sequence of 31 amino acids (S565-K595) in domain 6 of the light chain of high molecular weight kininogen (HK) has previously been shown to be responsible for the binding of plasma prekallikrein (PK) or kallikrein. To find effective peptides that might block binding between HK and PK on cell surfaces, a new series of synthetic peptides has now been prepared that incorporates portions of this binding domain sequence. For mapping the minimal sequence within HK, these new peptides were tested for their ability to compete with HK for binding PK in a cell-free system and on human umbilical vein endothelial cells (HUVEC). In the former, at pH 7.4, the kds for binding between kallikrein and either D567-K595, S565-P594, D567-S593, or D567-T591 were all similar to that for the binding of S565-K595 (0.2 to 0.4 micromol/L), but those for the binding of D568-K595, W569-K595, and D567-P589 were an order of magnitude greater (kd = 2 to 5 micromol/L). D567-S586, the shortest chain length of the N- and C-terminal truncation sequences tested, does not effectively compete with kininogen for kallikrein binding (kd = 100 micromol/L). These results imply that D567-T591, a 25-residue peptide (HK25c), contains sufficient structural information for binding kallikrein in solution. D567-T591 also is the minimum structural sequence to block binding of kallikrein to HUVEC-bound HK (IC50 = 50 nmol/L) and to inhibit PK activation to kallikrein on the cell surface (IC50 = 80 nmol/ L). In addition, D567-T591 also inhibits the generation of kallikrein-activated urokinase, which activates plasminogen to plasmin (IC50 = 100 nmol/L). Thus, HK-derived peptides may be useful compounds for modulating excessive fibrinolysis and hypotension in sepsis and multiple trauma.
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PMID:High molecular weight kininogen peptides inhibit the formation of kallikrein on endothelial cell surfaces and subsequent urokinase-dependent plasmin formation. 922 69

Recombinant human gamma interferon was used to treat 10 atopic dermatitis patients. Recombinant gamma interferon was administered weekly for three consecutive days at 50 microg/M2 SQ for four weeks. All patients' dermatitis improved with recombinant gamma interferon therapy and plasma tumor necrosis factor-alpha levels rose with treatment. Recombinant gamma interferon treatment positively correlated with reduced total plasma fibrinolysis as measured by the fibrin lysis plate, plasmin-alpha2antiplasmin complexes, and tissue type plasminogen activator levels. Accordingly, plasminogen activator inhibitor levels increased. Treatment also was associated with a transient increase in thrombin-antithrombin III complexes. Recombinant gamma interferon resulted in a significant increase in C1 inhibitor antigen but not activity. Plasma prekallikrein, high molecular weight kininogen, and factor XII levels were not decreased. However, 5 of the 10 atopic dermatitis patients before therapy had circulating cleaved plasma high molecular weight kininogen detected on immunoblot, indicating prior kallikrein formation. The cleaved, circulating plasma high molecular weight kininogen disappeared in four out of the five original patients who were reexamined at one year after treatment. These combined data indicated that recombinant gamma interferon treatment reduced total plasma fibrinolysis. In untreated atopic dermatitis, circulating cleaved high molecular weight kininogen also may be a presenting manifestation.
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PMID:Gamma interferon administration to patients with atopic dermatitis inhibits fibrinolysis and elevates C1 inhibitor. 966 47


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