Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.4 (trypsin)
 42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A large family has been studied, 11 of whose members have half-normal plasma concentrations of biological prothrombin activity. The pattern of inheritance is autosomal. By use of a specific immunoassay, affected family members have been shown to possess normal quantities of immunoreactive prothrombin, whose immunologic properties seem identical with those of the normal zymogen. Prothrombin isolation from the plasma of one such individual gave normal yields of protein but half-normal amounts of prothrombin activity. Activation of this material in the "intrinsic" and "extrinsic" systems, in concentrated sodium citrate, or by trypsin, gives rise to half, or less, of the thrombin clotting and esterase activities expected from a comparable normal prothrombin preparation. During the clotting of blood from an affected individual, all material with the mobility of prothrombin disappears. Immunoelectrophoresis of the serum reveals a normal nonthrombin "pro piece," and an additional activation product with an electrophoretic mobility intermediate between that of prothrombin and of "pro piece." These results suggest that affected individuals are heterozygotes in whom half the prothrombin molecules synthesized are structurally abnormal, since they undergo some alterations during activation, but are incapable of releasing the active enzyme, thrombin.
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PMID:Congenital dysprothrombinemia: an inherited structural disorder of human prothrombin. 535 38

The regulation of human Factor IXa was studied in vitro in human and mouse plasma and in vivo in the mouse. In human plasma, approximately 60% of the 125I-Factor IXa was bound to antithrombin III (ATIII) by 2 h, with no binding to alpha 2-macroglobulin or alpha 1-proteinase inhibitor, as assessed by gel electrophoresis and IgG- antiproteinase inhibitor-Sepharose beads. In the presence of heparin, virtually 100% of the 125I-Factor IXa was bound to ATIII by 1 min. The distribution of 125I-Factor IXa in mouse plasma was similar. The clearance of 125I-Factor IXa was rapid (50% clearance in 2 min) and biphasic and was inhibited by large molar excesses of ATIII-thrombin and alpha 1-proteinase inhibitor-trypsin, but not alpha 2-macro-globulin-trypsin; it was also inhibited by large molar excesses of diisopropylphosphoryl - (DIP-) Factor Xa, DIP-thrombin, and Factor IX, but not by prothrombin or Factor X. The clearance of Factor IX was also rapid (50% clearance in 2.5 min) and was inhibited by a large molar excess of Factor IX, but not by large molar excesses of Factor X, prothrombin, DIP-Factor Xa, or DIP-thrombin. Electrophoresis and IgG- antiproteinase inhibitor-Sepharose bead studies confirmed that by 2 min after injection into the murine circulation, 60% of the 125I-Factor IXa was bound to ATIII. Organ distribution studies with 125I-Factor IXa demonstrated that most of the radioactivity was in the liver. These studies suggest that Factor IXa binds to at least two classes of binding sites on endothelial cells. One site apparently recognizes both Factors IX and IXa, but not Factor X, Factor Xa, prothrombin, or thrombin. The other site recognizes thrombin, Factor Xa, and Factor IXa, but not the zymogen forms of these clotting factors. After this binding, Factor IXa is bound to ATIII and the complex is cleared from the circulation by hepatocytes.
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PMID:Regulation of factor IXa in vitro in human and mouse plasma and in vivo in the mouse. Role of the endothelium and the plasma proteinase inhibitors. 620 16

Vitamin K dependent carboxylation of an exogenous peptide substrate and endogenous protein substrates, vitamin K epoxidation, and reduction of vitamin K epoxide were measured in subcellular fractions from rat liver. The rough microsomal fraction was highly enriched in all four activities; lower levels were found in smooth microsomes. Mitochondria, nuclei, and cytosol had negligible activities. The addition of 0.2% Triton X-100 to intact microsomes resulted in a 10-20-fold stimulation in carboxylation of a peptide substrate. This marked latency suggests that the active site of the carboxylase may be accessible only from the lumen of the microsomal membrane. A lumen-facing orientation of the carboxylase was also supported by its inaccessibility to trypsin in intact microsomes contrasted with marked inhibition by trypsin in detergent-permeabilized microsomes. Vitamin K epoxidase and epoxide reductase activities were also inhibited by trypsin much more effectively in permeabilized than in intact microsomes, although some degree of exposure at the cytosolic surface was also indicated. These data suggest that carboxylation is an early event in prothrombin synthesis occurring primarily on the lumen side of the rough endoplasmic reticulum membrane. The location of the vitamin K epoxidation-reduction cycle enzymes is consistent with their possible role in the carboxylation reaction.
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PMID:Vitamin K dependent carboxylase: subcellular location of the carboxylase and enzymes involved in vitamin K metabolism in rat liver. 624 80

Coagulopathy is a hallmark of severe ferrous sulfate poisoning in humans and laboratory animals. Although nontransferrin-bound Fe3+ is thought to initiate the disorder, little is known about how it interferes with blood coagulation. At iron concentrations comparable to those of previous animal investigations, we reproduced the coagulopathy, in other words, the dose-related prolongation of the prothrombin, thrombin, and partial thromboplastin time, in human plasma in vitro. Studies of the mechanism by which iron prevents a normal plasma coagulation revealed that the proenzymes of the coagulation cascade and fibrinogen were not damaged by iron. Fibrinogen coagulability and fibrin monomer aggregation were unaffected by very high iron concentrations. Instead, thrombin was markedly inhibited by iron in its clotting effect on fibrinogen and, specifically, in its fibrinopeptide A-generating capacity, the inhibitory effect being reversible upon iron removal by EDTA chelation and gel filtration. Thrombin generation in the presence of iron was reduced as well, indicating an inhibition of one or several other enzymes of the intrinsic coagulation cascade. Because the amidolytic activity of human thrombin as well as factor Xa, kallikrein, and bovine trypsin was also reversibly suppressed by ferrous sulfate as well as ferric citrate, we consider it likely that the coagulopathy occurring in iron poisoning is the consequence of a general, physiologically important phenomenon: the susceptibility of serine proteases to nontransferrin-bound Fe3+.
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PMID:Blood coagulation and acute iron toxicity. Reversible iron-induced inactivation of serine proteases in vitro. 642 70

The entire amino acid sequence of complement factor B has been established combining both protein and DNA sequencing strategies. The zymogen consists of 739 amino acids, has four asparagine-linked carbohydrate sites, and has independently disulfide-bonded NH2- and COOH-terminal regions. The catalytic subunit, Bb, is a unique serine protease containing 259 amino acids that are not integral to any of the classical serine proteases. It is proposed that this region of the Bb fragment functions as a cofactor-binding domain for C3b. The Ba fragment was found to contain three regions of internal sequence homology which were unrelated to the "kringle" regions of prothrombin and plasminogen and which suggest an independent evolution for the B genome. Sequence alignment of the active site of B to the serine proteases was made using the three-dimensional structures of chymotrypsin and trypsin as molecular models. Three stretches within the hypothetical model for B contrast markedly with all known serine proteases in both amino acid sequences and predicted configuration. It is suggested that these "altered" regions contribute at least in part to the formation of the catalytic region of the C3 convertase.
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PMID:Complete primary structure for the zymogen of human complement factor B. 654 54

Bovine testicular hyaluronidase from various commercial sources showed the presence of an inhibitor of human plasma prothrombin time (PT). A testicular anticoagulant protein (TAP) was isolated from it by a 3-step procedure. The material was first passed through conconavalin-A-sepharose affinity chromatography where the anticoagulant material was separated from the hyaluronidase and protease which were retained by the column. In the second step the lower molecular weight proteins were removed by ultrafiltration. The supernatant which contained the anticoagulant protein was passed through the carboxymethyl cellulose column and the active material was eluted by 0.4 M NaCl solution. Sodium dodecyl sulfate (SDS) gel electrophoresis gave a molecular weight of approximately 35000. Unlike many small molecular weight proteins from bovine testes, TAP looses its anticoagulant property by heating for 30 minutes at 55 degrees C or by storage at pH 3.0 for 2 hours and it does not inhibit trypsin or thrombin. Its isoelectric pH was 9.7. Diisopropyl fluorophosphate (DFP) treated TAP was not effective as an anticoagulant.
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PMID:Isolation and properties of a new anticoagulant protein from commercial bovine testicular hyaluronidase. 661 84

The coagulation changes observed in acute pancreatitis were studied clinically and those 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 observed in six among 37 patients with acute pancreatitis and half of them died. Those changes observed clinically were either hypercoagulability or hypocoagulability. The coagulation changes after trypsin-induced acute experimental pancreatitis, elastase and autologous bile showed an indication of consumption coagulopathy. The effect upon blood coagulation after the intravenous injection of pancreatic juice included decreased platelet counts and plasma fibrinogen levels, prolonged partial thromboplastin and prothrombin time. The intravenous injection of pancreatic exudate produced greater changes than did those of an equal amounts of pancreatic juice. There was a shortening of E.L.T. and a marked increase in F.D.P. pancreatic exudate which accumulated during acute pancreatitis may contains a toxic substance or substances which contribute to the consumption of coagulation factors.
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PMID:[Disseminated intravenous coagulation in acute necrotizing pancreatitis]. 667 61

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.
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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.
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PMID:The nature of the binding for prothrombinase at the platelet surface as revealed by lipolytic enzymes. 706 May 71

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.
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PMID:[Effect of proteolysis inhibitors from Ascaris lumbricoides on the coagulation and fibrinolysis of human plasma]. 712 86


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