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)

The cyanogen bromide fragment CB67-129 of human prethrombin 1, corresponding to residues 54-116 of the thrombin B chain, is a potent chemotaxin for human peripheral blood monocytes and the murine macrophage like cell line, J774. Both of these cell types have been shown to respond chemotactically to alpha-thrombin and iPr2P-alpha-thrombin. Effective concentrations for stimulating directed cell movement with the fragment vary from 10(-11) to 10(-7) M. Moreover, CB67-129 and its parent protein compete for the same chemotactic receptor site. Fragment CB67-129, representing residues 54-116 of the human thrombin B chain sequence, contains a nine-residue insertion ("loop B") that is absent in homologous sequences derived from the closely related proteases chymotrypsin and trypsin. Unlike iPr2P-alpha-thrombin, iPr2P derivatives of these latter enzymes possess little or no chemotactic activity, suggesting a relationship between the insertion sequence and thrombin chemotactic activity. The loop B sequence is unique insofar as it contains all of the carbohydrate moieties known to reside in alpha-thrombin. However, chemotactic activity is only minimally reduced subsequent to hydrolysis by both neuraminidase and beta-galactosidase, indicating that receptor recognition and stimulated cell movement are mainly a function of structure of the cyanogen bromide derived fragment rather than of asparagine-linked carbohydrates.
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PMID:Localization of a chemotactic domain in human thrombin. 670 77

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

Heparin accelerates the rate of inhibition of thrombin by antithrombin III. Reduction of one of the three antithrombin disulfide bonds with dithiothreitol under mild conditions abolishes this rate-enhancing effect without affecting the rate of reaction in the absence of heparin. Alkylation of mildly reduced antithrombin III with [3H]iodacetic acid followed by digestion with cyanogen bromide yielded two major labeled peptides. The smaller peptide, containing Cys-422, was identified as extending from Gly-414 to the C-terminus, Lys-424. Our data are consistent with the larger labeled peptide being the one extending from Glu-104 to Met-243 and containing Cys-239. Cys-422 has been shown by others to be linked to Cys-239. These data indicate that the sensitive disulfide bond in antithrombin III extends between Cys-239 and Cys-422; the site at which thrombin cleaves the antithrombin III is between these two half-cystines.
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PMID:Localization of the disulfide bond in human antithrombin III required for heparin-accelerated thrombin inactivation. 683 Feb 63

Using DEAE-Sephadex A-50 column chromatography and gel filtration, a potent platelet aggregation inhibitor from Trimeresurus gramineus venom was purified. It was an acidic phospholipase A, rich in aspartic acid, glutamic acid and half-cystine, with an isoelectric point of 3.6. At a concentration of 10 micrograms/ml, the purified inhibitor showed a marked inhibitory effect on platelet aggregations induced by adenosine diphosphate, collagen, sodium arachidonate and ionophore A-23187 in rabbit platelet-rich plasma, washed platelet suspension, as well as in thrombin-degranulated platelet suspension. The ID50 of this venom inhibitor was about 2.5-5 micrograms/ml in platelet aggregations induced by all these aggregation inducers. The action of this inhibitor could be partially antagonized by phosphatidylethanolamine. High concentration of Ca2+ (5 mM) did not reverse the inhibitory action even in the presence of ionophore A-23187. The [14C]serotonin release induced by sodium arachidonate and thrombin was unaffected. Malonic dialdehyde formation induced by these aggregation inducers remained unchanged. Basal and prostaglandin E1-stimulated cAMP levels were not altered by this inhibitor. No lactate dehydrogenase was released even at a concentration of 62.5 micrograms/ml. Polylysine-induced platelet agglutination was not affected. beta-Mercaptoethanol inactivated both its phospholipase A enzymatic and platelet inhibitory activities, while p-bromophenacyl bromide only inactivated the former activity. The possibility of acting on a common final step of platelet aggregation, i.e. the intercellular adhesion between the activated platelets, was proposed.
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PMID:Potent platelet aggregation inhibitor from Trimeresurus gramineus snake venom. 684 80

Chemical modification studies have demonstrated that the ultra-violet difference spectrum of alpha-thrombin produced in the presence of sodium is due primarily to changes in the environment of tyrosine residues. This is based on the observation that the spectrum could be abolished by treatment of alpha-thrombin with tetranitromethane but not with dimethyl-(2-hydroxy-5-nitrobenzyl) sulfonium bromide. Although lithium produces similar (UV) difference spectrum, circular dichroism studies indicate that sodium and lithium induce different conformational transitions. alpha-Thrombin tends to assume a more ordered structure in the presence of sodium whereas lithium has the reverse effect. This inverse behavior is consistent with the effects of these cations on the autolysis rate and thermal stability of the activities of alpha-thrombin.
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PMID:Effects of sodium and lithium salts on the conformation of human alpha-thrombin. 685 97

Heparin kinetics were determined in four normal subjects, each of whom received three different doses (25, 50, and 75 units/kg body weight) by intravenous injection. Multiple blood samples were collected after each dose and each plasma sample was assayed for heparin activity using three different assay methods. Two of these assays are based on coagulation tests, i.e., activated partial thromboplastin time and thrombin time, while the third is based on chemical neutralization of heparin using hexadimethrine bromide. Heparin kinetics showed pronounced dose-dependent changes, irrespective of the assay method used, which were characterized by increasing biologic half-life and decreasing total clearance (Cl) with increasing dose. No changes were noted in apparent volume of distribution (Vd). This data also showed that there were differences in kinetic parameters of heparin depending on the assay method. In general, values for total Cl and apparent Vd based on chemical neutralization were approximately 1.5 to 2.0 times these parameters based on coagulation tests. We conclude that the immediate mechanism of the dose-dependent heparin kinetics is decreasing total clearance with increasing dose and suggest that in vivo activation of the anticoagulant properties of heparin may explain the assay-dependent kinetics.
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PMID:Heparin kinetics determined by three assay methods. 705 98

The amino acid sequence of the cyanogen bromide (CNBr) fragment extending from position 24 to position 98 of the H-2Dd murine histocompatibility antigen has been determined by using radiochemical microsequencing techniques. This 75-residue fragment (CN-C) is one of two glycopeptides generated by CNBr cleavage of the extracellular portion of the H-2Dd molecule (H-2Dd papain). Determination of this sequence completes the amino-terminal 100 residues of the H-2Dd molecule. The primary structure of CN-C was established by thrombin digestion of isolated CN-C and sequence determination of the three resulting peptides. The COOH-terminal met and its adjacent residue were determined by sequence analysis of a tryptic peptide which overlaps CNBr fragments C and b4 (residues 99-138). Alignment of the thrombic peptides was accomplished by NH2-terminal sequence analyses of CN-C and peptides generated by Staphylococcus aureus V8 protease digestion of CN-C. The sequence data presented here, together with that already given for H-2Dd [Nairn, R., Nathenson, S. G., & Coligan, J.E. (1980) Eur. J. Immunol. 10,495-503], allow a comparison of the NH2-terminal 100 residues of the Dd, Db, Kd, and Kb molecules. Discrete areas of diversity, in particular, one between residues 62 and 83, are obvious. Comparison over some 180 residues of the Dd and Kb molecules reveals a particularly close similarity between these products of a K and a D gene from widely disparate mouse strains.
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PMID:Amino acid sequence of cyanogen bromide fragment CN-C (residues 24-98) of the mouse histocompatibility antigen H-2Dd. A comparison of the amino-terminal 100 residues of H-2Dd, Dd, Kd, and Kb reveals discrete areas of diversity. 729 46

The conformations of human alpha-thrombin and gamma-thrombin have been compared by circular dichroism, solvent perturbation different spectroscopy, and chemical modification. Circular dichroism studies indicate that proteolytic conversion of alpha-thrombin to gamma-thrombin is accompanied by considerable conformational changes which include a decrease in alpha-helical content from 5-7% to 0-1%. Solvent perturbation at pH 6.0 obtained with 20% ethylene glycol, 20% glycerol, and 20% dimethyl sulfoxide indicates an apparent exposure of 3.5 +2- 0.2 tryptophan and 7.8 +/- 0.1 tyrosine residues in alpha-thrombin and 4.6 +/- 0.2 tryptophan and 9.2 +/0 0.3 tyrosine residues in gamma-thrombin. This increased exposure is substantiated by the greater reactivity of tryptophan residues in gamma-thrombin toward dimethyl (2-hydroxy-5-nitrobenzyl) sulfonium bromide. It suggests that gamma-thrombin is a less compact molecule than the parent alpha-thrombin. Solvent perturbation studies of alpha-thrombin and gamma=thrombin inhibited by phenyl-methanesulfonyl fluoride showed that 0.3 +/- tryptophan and 0.9 +/- 0.3 tyrosine residues in alpha-thrombin and 0.6 +/- 0.3 tryptophan and 1.3 +/- 0.4 tyrosine residues in gamma-thrombin were blocked by the inhibitor. These subtle differences in the extent of blocking of tyrosine and tryptophan suggest a tighter conformation in the catalytic site of gamma-thrombin compared to that of alpha-thrombin.
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PMID:Conformational differences between high clotting human alpha-thrombin and nonclotting gamma-thrombin. 730 21

Chemical modification of antithrombin III, the major plasma protease inhibitor, with the tryptophan reagent dimethy(2-hydroxy-5-nitrobenzyl) sulfonium bromide, results in the incorporation of one hydroxynitrobenzyl moiety per molecule of antithrombin III. The derivatized inhibitor does not exhibit the heparin-promoted enhancement in rate of thrombin inactivation which is characteristic of the native molecule. However, the rates of thrombin inactivation in the absence of heparin are identical with native and derivatized inhibitors, indicating that the site of protease . inhibitor complex formation is not altered. Unlike native antithrombin III, the modified inhibitor does not bind to a heparin-agarose affinity column. When the modification reaction was performed with added heparin, the extent of modification was decreased and the heparin-promoted enhancement of thrombin inactivation was preserved. These results indicate that the integrity of a specific tryptophan residue is critical to the binding of heparin to antithrombin III.
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PMID:The heparin binding site of antithrombin III. Evidence for a critical tryptophan residue. 735 60

An approach to providing anticoagulant activity to biomedical materials was presented, applying an immobilization technique. Antithrombin III (AT III) inactivates the activated coagulation factors including Factor Xa and thrombin. Heparin stimulates the inactivation of Factor Xa and thrombin by AT III. Thus AT III and heparin were co-immobilized on Sepharose 4B, polyvinyl alcohol, polyhydroxy-ethyl methacrylate and silicone-coated nylon by the cyanogen bromide procedure. Those co-immobilized preparations, abbreviated as I-AT III . Hep, actively neutralized both Factor Xa and thrombin. The activity of I-AT III . Hep was much higher than immobilized heparin and/or immobilized AT III. I-AT III . Hep, like soluble AT III and heparin, instantaneously neutralized both thrombin and Factor Xa. When two enzymes, thrombin and Factor Xa, were present, I-AT III . Hep neutralized Factor Xa in preference to thrombin : The neutralization of thrombin was inhibited by the presence of Factor Xa, but neutralization of Factor Xa was independent of the presence of thrombin. The amount of Factor Xa neutralized was higher than that of thrombin.
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PMID:Anticoagulant activity of artificial biomedical materials with co-immobilized antithrombin III and heparin. 741 94


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