Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.6 (thromboplastin)
 13,278 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Mouse placental cells have been isolated and grown in cultures. These cells produce a procoagulant which is identical to thromboplastin (factor III) by two criteria. The procoagulant activity increases with time, culminating on the 5th day of culture. The increase is inhibited by cycloheximide, alpha-amanitin, and actinomycin D, showing that de novo synthesis of protein and RNA is necessary. About 90-95% of the total cellular thromboplastin activity in whole cells is available for inactivation by particle-bound trypsin and thus present on the cell surface. Endotoxin and phytohaemagglutinin did not further increase the procoagulant activity.
 ...
PMID:Synthesis of thromboplastin (factor III) in mouse placental cells in vitro. 687 49

The subsite specificities of bovine factor IXa, factor Xa, factor XIa, factor XIIa, thrombin, plasma kallikrein, and trypsin were mapped with amino acid, dipeptide, and longer peptide thioester substrates. Each substrate contained a P1 Arg residue. The P1' residues included thiol residues which are analogues of valine, leucine, and isoleucine, respectively, and the P2 residue included 12 representative amino acid residues. Longer substrates with the sequence at the antithrombin III reactive site and at the zymogen activation site of various coagulation factors were also studied. The enzymatic hydrolysis of the thioesters was measured in the presence of 4,4'-dithiodipyridine which provides a very sensitive assay for the free thiol. The thioesters were excellent substrates for the coagulation factors studied, and the kcat/Km values for the best thioester substrates were higher than those previously reported for most of these enzymes. Thrombin and plasma kallikrein were the most active of the coagulation factors toward the thioester substrates. The best substrate for thrombin was Z-Gly-Arg-SCH2C6H5, although substrates containing proline in the P2 position were also quite effective. Some of the better substrates for plasma kallikrein had a P2 Phe or Trp residue. Factor IXa was the least reactive of the coagulation factors and hydrolyzed only four of the dipeptide thioesters. Substrates with bulky hydrophobic groups such as Phe or Trp in the P2 position were the most reactive with factor IXa. Factor Xa hydrolyzed all the thioester substrates tested, the most reactive being Z-Gly-Arg-SCH2C6H5. This is consistent with the fact that glycine and arginine are present in the P2 and P1 positions, respectively, of the factor Xa sensitive bonds in prothrombin which is the physiological substrate for factor Xa. Bovine factor XIa showed the least amount of specificity of the various coagulation factors and was quite reactive toward all of the thioester substrates. The most sensitive substrate for this enzyme was also Z-Gly-Arg-SCH2C6H5. Factor XIIa preferred the dipeptide with a P2 Phe, although the simpler thioester Z-Arg-SCH2CH(CH3)2 was more reactive. Trypsin hydrolyzed all of the thioester substrates at a high rate and showed little substrate specificity. With all enzymes studied, extension of the thioester substrate beyond P2 or the P1' thiol leaving group did not lead to an improvement in hydrolysis. Due to their high kcat/Km values and the ease of detecting the thiol leaving group, thioester substrates should be extremely useful for future studies of coagulation proteases.
 ...
PMID:Mapping the active sites of bovine thrombin, factor IXa, factor Xa, factor XIa, factor XIIa, plasma kallikrein, and trypsin with amino acid and peptide thioesters: development of new sensitive substrates. 697 85

The nature of the receptor for the prothrombinase complex at the surface of non-activated platelets was investigated by measuring the platelet prothrombin-converting activity wih a chromogenic substrate assay, after treatment of the platelets with various phospholipases or three different proteolytic enzymes. Platelet prothrombin-converting activity only decreased after treatment with those phospholipases which are able to hydrolyse phospholipids in the intact platelet and also have the ability to degrade negatively charged phospholipids, phosphatidylserine and phosphatidylinositol. Those phospholipases which do hydrolyse phospholipids in the intact platelet but have no activity towards phosphatidylserine (and phosphatidylinositol) produce an increase in the platelet prothrombin-converting activity. Proteolytic treatment of platelets with trypsin, chymotrypsin or papain did not result in a decrease of prothrombin-converting activity. It is concluded that negatively charged phosphatidylserine and possibly phosphatidylinositol are involved in the prothrombin-converting activity of non-activated platelets. We could not demonstrate the involvement of platelet membrane proteins in a receptor for the components of the prothrombinase complex.
 ...
PMID:The nature of the binding for prothrombinase at the platelet surface as revealed by lipolytic enzymes. 706 May 71

Platelets are able to stimulate an increase in two distinct activities, tissue factor (thromboplastin) and fibrinolytic inhibition, in human fibroblasts in vitro. A procedure has been developed which allows the purification of a platelet macromolecule which is able to stimulate both of these changes. Washed human platelets were homogenized, sonicated, and then centrifuged at 90,000 x g for 2 h. The resulting pellet was solubilized in 0.05 M sodium carbonate, pH 10.5, and chromatographed on Sephadex G-200, then on hydroxylapatite, resulting in a 135-fold purification and a 20% yield. When the purified material was further fractionated on sodium dodecyl sulfate-polyacrylamide gels, stimulatory activity for both tissue factor and fibrinolytic inhibition was found only in the 75,000-dalton region. The active material could be inactivated by mercaptoethanol with no change in its apparent molecular weight. It was readily inactivated by trypsin with the concomitant loss of the 75,000-dalton Coomassie-staining band. Assay of the purified material for carbohydrate was negative. After isoelectric focusing, the purified material had a major band at pH 5.8 which stimulated both tissue factor and fibrinolytic inhibition. Subcellular fractionation of platelet homogenates by sucrose density gradient centrifugation resulted in a 2-fold increase in stimulatory material in the granule/mitochondrial fraction. This platelet-derived protein may represent a physiologically important regulator for both cellular procoagulant and the net fibrinolytic activity of systemic cells.
 ...
PMID:Purification of a platelet protein which stimulates fibrinolytic inhibition and tissue factor in human fibroblasts. 711 21

By means of the method of ROLA & PUDLES modified by the authors the effect of trypsin and chymotrypsin inhibitors isolated from body walls of Ascaris lumbricoides on coagulation and fibrinolysis of human plasma in vitro was studied. The effect of both Ascaris inhibitors on coagulation phases I and II, the inhibition of thromboplastin and thrombin generations (thromboelastography, prothrombin and thrombin times) and the fibrinogenesis retardation of human plasma (time of lysis of euglobulin clot, time of clot fibrinolysis activated by streptokinase) were found. "The stairs phenomenon" was observed on thromboelastographic curves. The plasmin activity in an active form as well as its formation from plasminogen by streptokinase activation were reduced by chymotrypsin and trypsin Ascaris inhibitors alike.
 ...
PMID:[Effect of proteolysis inhibitors from Ascaris lumbricoides on the coagulation and fibrinolysis of human plasma]. 712 86

The coagulation changes observed in acute experimental pancreatitis were compared with those after the intravenous infusion of pancreatic juice and ascitic fluid exudate obtained from bile-induced pancreatitis in dogs. The coagulation changes after acute pancreatitis was induced by the intraductal injection of autologous bile, trypsin or elastase showed decreased platelet counts, decreased plasma fibrinogen levels, prolonged partial thromboplastin and prothrombin times, shortened euglobulin clot lysis time and increased fibrin degradation products. Multiple microemboli were observed in the lung and, occasionally, in the kidney, an indication of consumption coagulopathy. The effects upon blood coagulation after the intravenous injection of pancreatic juice included decreased platelet counts, decreased plasma fibrinogen levels and prolonged partial thromboplastin and prothrombin times. The intravenous injection of pancreatic exudate produced greater changes than did those of an equal amount of pancreatic juice. There was a shortening of euglobulin clot lysis time and a marked increase in fibrin degradation products. Pancreatic exudate which accumulates during acute pancreatitis may contain a toxic substance or substances which contribute to the consumption of coagulation factors during acute pancreatitis.
 ...
PMID:The effects upon blood coagulation in dogs of experimentally induced pancreatitis and the infusion of pancreatic juice. 726 8

The tick Boophilus microplus contains a protein that inhibits a range of proteolytic enzymes. Variations in the concentration of this protein throughtout the life cycle were followed by measuring simultaneously the inhibition of trypsin and chymotrypsin and reaction with an antiserum to the purified inhibitor. The protein is present in large amounts in eggs and in unfed larvae, but its concentration falls very rapidly after the start of the parasitic stage of the life cycle. This, together with previous evidence, suggests that the inhibitor is important both in eggs and in the initial establishment of the parasite on its host. The activity of the protein towards several enzymes has been measured as an indication of its possible function. Bovine trypsin, chymotrypsin and plasmin and pig pancreatic kallikrein are all inhibited. The protein also affects the blood-coagulation system at several points, since it prolongs both activated-partial-thromboplastin time and prothrombin time. It inhibits the complement-dependent lysis of erythrocytes, but is without significant effect on mitogen-induced lymphocyte stimulation. Thus the inhibitor could have several effects on the host that would be beneficial to the parasite.
 ...
PMID:On the biological role of a proteolytic-enzyme inhibitor from the ectoparasitic tick Boophilus microplus. 745 13

Crystal structures of DX9065a and a related bisamidino-aryl inhibitor specific for the blood-clotting factor Xa have been solved in complex with bovine beta-trypsin to a resolution of 1.9 A. Each inhibitor exhibits an extended conformation along the active site, in contrast to the compact folded structures observed for thrombin specific inhibitors. Few direct contacts (predominantly in the S1 pocket) are made between trypsin and the inhibitors. Transfer of the inhibitors to the active site of factor Xa suggests a three-site interaction: salt bridge formation at the base of the primary specificity pocket, extensive hydrophobic surface burial and a weak electrostatic interaction between the distal basic component of the inhibitor and an electronegative cavity of factor Xa formed by three backbone carbonyl oxygens. Additivity of these three interactions is the basis for the observed strong inhibition of factor Xa and provides a framework for the design of novel factor Xa inhibitors. A propionic acid group of the inhibitor would clash with the thrombin specific '60-insertion loop', thus conferring selectivity against thrombin.
 ...
PMID:Crystal structures of factor Xa specific inhibitors in complex with trypsin: structural grounds for inhibition of factor Xa and selectivity against thrombin. 749 54

Triabin, a new thrombin inhibitor, has been purified from the saliva of Triatoma pallidipennis, a blood-sucking triatomine bug. It forms a noncovalent complex with thrombin at a molar ratio of 1:1, inhibits thrombin-induced platelet aggregation, and prolongs thrombin clotting time and activated partial thromboplastin time. However, it only minimally suppresses the amidolytic activity of thrombin, as measured by a chromogenic peptide substrate assay. It completely blocks trypsin-catalyzed cleavage of thrombin, probably via protection of the anion-binding exosite and inhibits the effect of thrombomodulin on thrombin in a dose-dependent fashion. These results indicate that the inhibitor is directed toward the anion-binding exosite of thrombin. The protein was partially sequenced and the information used to isolate cDNA clones from a T. pallidipennis salivary gland library. Four slightly polymorphic variants coding for mature proteins of 142 amino acids preceded by a putative leader sequence were obtained. The recombinant protein expressed in the periplasmic space of Escherichia coli has a biological activity similar to that of salivary triabin, as tested in a thrombin-induced platelet aggregation assay. In addition, recombinant triabin inhibits thrombin-catalyzed hydrolysis of fibrinogen with a Ki of about 3 pM.
 ...
PMID:Triabin, a highly potent exosite inhibitor of thrombin. 749 80

The human intracellular serine proteinase inhibitor, proteinase inhibitor 6 (PI-6), was expressed in the methylotropic yeast Pichia pastoris. The PI-6 cDNA was modified to encode six histidine residues immediately after the initiation codon, and was placed under the control of the P. pastoris alcohol oxidase promoter in the vector pHIL-D2. On the methanol induction, active recombinant PI-6 was produced within the yeast cells, and following cell lysis, was separated from yeast proteins by affinity chromatography using nickel nitrilo-tri-acetic acid (NTA) resin. The interaction of recombinant PI-6 with a range of serine proteinases was studied. Second order association rate constants (ka) were derived for the interaction with trypsin (1.8 x 10(6) M-1 s-1), thrombin (1.2 x 10(5) M-1 s-1), urokinase plasminogen activator (4.0 x 10(4) M-1 s-1), plasmin (1.3 x 10(6) M-1 s-1), and activated protein C (7.5 x 10(3) M-1 s-1). By monitoring complex formation, recombinant PI-6 was also shown to interact with factor Xa. No complex formation was observed with chymotrypsin, human leukocyte elastase, cathepsin G and tissue plasminogen activator, although PI-6 is apparently a substrate for chymotrypsin, leukocyte elastase and cathepsin G.
 ...
PMID:Production and characterization of recombinant human proteinase inhibitor 6 expressed in Pichia pastoris. 754 63


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>