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)

A metalloprotease from the rattlesnake Crotalus atrox venom was isolated and purified from multiple-step chromatographies including anion-exchange chromatography, gel permeation and reversed-phase HPLC. The fraction was shown to be homogeneous as judged by SDS-gel electrophoresis. It also showed a high proteolytic activity against alpha- and beta-chains of fibrinogen molecules. Further characterization of the purified fraction with fibrinogenase activity indicated that it is a single-chain protease with a molecular mass of about 24 kDa and an acidic isoelectric point. It is relatively heat stable up to about 65 degrees C, inhibited by EDTA, beta-mercaptoethanol, but not by phenylmethanesulfonyl fluoride, N alpha-p-tosyl-L-phenylalanine chloromethyl ketone and N alpha-p-tosyl-L-lysine chloromethyl ketone, soybean trypsin inhibitor and aprotinin. Amino acid analysis showed that the enzyme possesses an amino acid composition very similar to some metalloproteases characterized before from the closely related rattlesnake venoms. N-Terminal sequence analysis of the enzyme corroborated some similarity between this enzyme and the reported sequences of these enzymes characterized from the Crotalidae snake family. This study indicated the presence of a novel fibrinogenase (termed Catroxase) with N-terminal sequence different from the metalloprotease with hemorrhagic activity isolated from the same Western diamondback rattlesnake.
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PMID:Characterization of a protease with alpha- and beta-fibrinogenase activity from the Western diamondback rattlesnake, Crotalus atrox. 152 Mar 24

Fibrolase, a fibrinolytic enzyme isolated from Agkistrodon c. contortrix (southern copperhead) venom, solubilizes fibrin primarily by rapid hydrolysis of the alpha and beta chains. Fibrolase is also an A alpha, B beta fibrinogenase. The breakdown products of fibrin and fibrinogen following incubation with fibrolase were different from those observed with plasmin. This enzyme is a metalloprotease that was inhibited by ethylenediaminetetraacetic acid. Fibrolase was inhibited by dithiothreitol, suggesting that disulfide bonds are important for catalytic activity. It was also inhibited by alpha 2-macroglobulin, but not by the soybean or lima bean trypsin inhibitors, diisopropylfluorophosphate, or p-hydroxymercuribenzoate. Unlike thrombolytic agents such as streptokinase, fibrolase does not activate plasminogen as evidenced by the use of plasmin-specific chromogenic substrate S-2251 and by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
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PMID:Biochemical characteristics of fibrolase, a fibrinolytic protease from snake venom. 169 22

Cultured guinea pig bone marrow megakaryocytes were found to secrete a 92-kd collagenase that was detected by digestion of gelatin in a polyacrylamide substrate gel assay. Neither casein or bovine serum albumin were digested by this enzyme. The enzyme is a neutral metalloprotease. Its secretion is increased by thrombin (1.0 U/ml) and phorbol myristate acetate (10 ng/ml) and is unaffected by prostaglandin E1 (10 microM). In the absence of serum, gelatinase secretion is inhibited, but it can be stimulated by cytochalasin D (1.0 microgram/ml). Gelatinase activity in the medium from megakaryocytes cultured on rat tail collagen gel is decreased. Medium from megakaryocytes cultured on Matrigel contains a second gelatinase of 90 kd. Addition of the tetrapeptide RGDS to the cultures on Matrigel blocks the appearance of the 90-kd gelatinase. Platelets contained both a 92- and a 90-kd gelatinase that was detected only after thrombin activation. The results indicate that megakaryocytes can secrete a collagenase, and its secretion may be in part controlled by interaction with the extracellular matrix. The appearance of the 90-kd gelatinase may be associated with megakaryocyte maturation and platelet formation.
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PMID:Collagenase production by guinea pig megakaryocytes in vitro. 216 10

Serine proteases in mast cell granules, such as chymase, atypical chymase, and tryptase, which are major proteins in the granules, may play important roles in the process of immunoglobulin E (IgE)-mediated degranulation and in pathobiological alterations in tissues. Indeed, inhibitors of chymase, substrate analogs, and antichymase F(ab')2, but not inhibitors of tryptase, markedly inhibited histamine release induced by IgE-receptor bridging but not that induced by Ca ionophore. In contrast, inhibitors of metalloprotease inhibited histamine release induced not only by IgE-receptor bridging but also by Ca ionophore. These results suggest that chymase and metalloprotease are involved at different steps in the process of degranulation. The extents of inhibition of histamine release were closely correlated with the amounts of the inhibitors of chymase accumulated in the granules. After degranulation, the released proteases may in part contribute to pathobiological alterations in allergic disorders through generations of C3a anaphylatoxin and thrombin by human and rat tryptase, respectively, and those of angiotensin II and a chemotactic factor of neutrophils by human and rat chymase, respectively. Moreover, chymase and atypical chymase from rat were shown to destroy type IV collagen, and human tryptase was found to hydrolyze various plasma proteins, such as fibrinogen and high-molecular-weight kininogen. The biological activities of tryptase and chymase from rat may be regulated by their dissociation from and association with trypstatin, an endogenous inhibitor of these proteases.
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PMID:Biological functions of serine proteases in mast cells in allergic inflammation. 246 15

The effect of a zinc metalloprotease from Serratia marcescens on platelet surface glycoproteins (GP) Ib and V was analyzed. Increasing protease treatments caused progressive loss of GP Ib with appearance of the major fragment, glycocalicin, in the supernatant solution. No GP V was detected in the supernatant solution, and protease-pretreated platelets had the same capacity as control platelets to release fragment 1 of GP V in response to thrombin. The Serratia protease-pretreated platelets did show the lag before thrombin-induced dense granule secretion, characteristic of platelets modified by pretreatment with other nonstimulating proteases. Treatment with Serratia protease gives the only demonstrated selective loss of GP Ib without apparent effect on GP V. It suggests that GP V (1) does not depend on GP Ib for its association with platelets and (2) is not the substrate for protease modification of platelet function.
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PMID:Differential effect of Serratia protease on platelet surface glycoproteins Ib and V. 315 41

Leech Haementeria ghilianii has an anticoagulant in its salivary glands that renders ingested blood incoagulable by thrombin. The mechanism of blood incoagulation is associated with cleavage of peptide bonds in fibrinogen, and thus the active agent, called hementin, is a proteolytic enzyme. Hementin was isolated and purified 16-fold from the anterior salivary glands by anion-exchange chromatography, ammonium sulfate fractionation, and cation exchange chromatography. Pure material obtained by slab gel electrophoresis contained a single polypeptide chain with approximate Mr of 120,000. Hementin was stable for many hours at room temperature, but on incubation at 60 degrees C for 15 min all activity was lost. At 4 degrees, hementin had no activity, but this inhibition was fully reversible. Complete inactivation occurred in the presence of EDTA, cysteine, DTT, sodium phosphate and at extreme pH (greater than 11 or less than 5), whereas citrate, Tris, glycine, and EGTA caused only partial loss of activity. DFP, PMSF, iodoacetic acid, and leupeptin had no effect on hementin activity. The data indicated hementin to be a neutral metalloprotease with optimum pH of 7.5. The enzyme had high affinity for cleaving fibrinogen and calculations of kinetic data from a double-reciprocal plot gave a Km of hementin for fibrinogen of 1.0 +/- 0.1 microM. Normal human citrated plasma or fresh blood were rendered incoagulable after incubation with hementin, indicating that the enzyme activity was not affected by plasma protease inhibitors. Plasma levels of coagulation factors II, V, VII, VIII, IX, X, XI, XII, prekallikrein, and high-molecular-weight kininogen were not altered by the enzyme. Hementin, a neutral metalloprotease resistant to plasma protease inhibitors, executes its anticoagulant effect on blood by selective cleavage of fibrinogen.
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PMID:Hementin: anticoagulant protease from the salivary gland of the leech Haementeria ghilianii. 636 Nov 87

Formation of thrombin during incubation of purified bovine prothrombin with purified staphylococcal metalloprotease has been investigated. Thrombin activity was estimated by examination of clotting time and by digestion of a synthetic substrate, Chromozym TH. The metalloprotease caused direct activation of prothrombin which was inhibited by the addition of ethylenediaminetetraacetic acid. Metalloprotease produced by some strains of Staphylococcus aureus may simulate staphylocoagulase activity.
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PMID:Prothrombin activation by a metalloprotease from Staphylococcus aureus. 678 2

A non-hemorrhagic metalloprotease (protease L4) was purified from the venom of Chinese Mamushi (Agkistrodon halys brevicaudus) by gel filtration and anion-exchange chromatography. Protease L4 has the molecular weight of 22,000 and its optimum pH was 8.5. The protein was stable in the pH range of 5-9 and below 40 degrees C. The proteolytic activity was inhibited by metal-chelating agents and some metal ions. Calcium ion activated the activity dose-dependently, but had only a minor effect on the thermal and pH stability. L4 showed fibrinogenase activity, hydrolyzing only the A alpha chain of fibrinogen. The protease cleaved preferentially at the N-terminal of Leu and His residues of some peptides.
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PMID:Purification and characterization of a non-hemorrhagic metalloprotease from Agkistrodon halys brevicaudus venom. 782 14

Kistomin, a metalloprotease purified from venom of Calloselasma rhodostoma, dose- and time-dependently prolonged the latent period of aggregation and inhibited ATP secretion of human washed platelets stimulated by thrombin. It inhibited aggregation induced by low concentrations of thrombin (< or = 0.2 U/ml) whereas it had only slight effect on aggregation induced by high concentrations of thrombin (> or = 0.5 U/ml). Meanwhile it also inhibited ristocetin-induced platelet aggregation in a dose- and time-dependent manner. It significantly inhibited cytosolic calcium rise of Quin 2--loaded platelets, completely blocked thromboxane B2 formation, and blocked [3H]inositol phosphates formation of [3H]myoinositol loaded platelets stimulated by 0.1 U/ml of thrombin. Kistomin inhibited significantly thromboxane but not [3H]inositol phosphates formation of platelets stimulated by a high concentration of thrombin (1 U/ml). Incubation of platelets with kistomin resulted in a selective cleavage of platelet membrane glycoprotein Ib as revealed by SDS/PAGE stained by periodic acid/Schiff reagent. These results suggested that thrombin activates platelets at least through two receptors/or effectors-mediated events. In addition to glycoprotein Ib, other surface membrane component(s) (e.g., the seven transmembrane domain thrombin receptor) may also be important in regulating the biochemical events of human platelets in response to thrombin. However, the extent and rate of platelet aggregation stimulated by low concentrations of thrombin ( < or = 0.2 U/ml) are closely related with the intactness of glycoprotein Ib.
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PMID:Antiplatelet protease, kistomin, selectively cleaves human platelet glycoprotein Ib. 825 30

The venom of the viper Echis carinatus contains a metalloprotease, ecarin, that is a potent prothrombin activator. We here show that the venom is also rich in another prothrombin activator, which does not belong to any known category of prothrombin activators. The novel enzyme, designated carinactivase-1 (CA-1), consists of two subunits held together non-covalently but very tightly. One subunit is a 62-kDa polypeptide that has metalloprotease activity and is homologous to the single-chain enzyme ecarin; the other subunit of 25 kDa consists of two disulfide-linked polypeptides of 17 and 14 kDa, and this subunit resembles the anticoagulant in the habu snake venom, IX/X-bp, that specifically binds the Gla domains of coagulation factors IX and X in a Ca2+-dependent fashion. The activation of prothrombin by CA-1 requires Ca2+ ions at millimolar concentrations and in the absence of Ca2+ ions this enzyme is virtually inactive. By contrast, activation by ecarin is completely independent of Ca2+ ions. CA-1, unlike ecarin, does not activate prothrombin derivatives, in which binding of Ca2+ ions has been perturbed, namely prethrombin-1 and acarboxyprothrombin. Furthermore, the isolated catalytic subunit, although its activity is greatly reduced as compared to that of the holoenzyme, no longer requires Ca2+ ions for the activation of prothrombin. Reconstitution with the non-catalytic 25-kDa subunit restores high level activity and the dependence on Ca2+ ions. Finally, prothrombin activation by CA-1 is inhibited by prothrombin fragment 1, and the isolated non-catalytic subunit is capable of binding fragment 1 in the presence of Ca2+ ions. From these observations, we postulate the following unique mechanism for the activation of prothrombin by CA-1. The enzyme primarily recognizes the Ca2+-bound conformation of the Gla domain in prothrombin via the 25-kDa regulatory subunit, and the subsequent conversion of prothrombin to active thrombin is catalyzed by the 62-kDa catalytic subunit.
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PMID:Isolation and characterization of carinactivase, a novel prothrombin activator in Echis carinatus venom with a unique catalytic mechanism. 861 3


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