Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.5 (thrombin)
 33,306 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A small fragment of C3, called C3a, which has smooth muscle contracting activity, was isolated by three different methods. At pH 8.6, C3a behaved as cation, and using the Archibald method, its mol wt was determined to be 7000. A specific antiserum to C3a showed the fragment to be antigenically distinct from the rest of the C3 molecule, i.e., the C3b portion. The same antiserum and an anti-whole C3 were able to inhibit the biologic activity of C3a. In addition to anaphylatoxin activity, leukocyte chemotactic activity was shown to reside in C3a. Treatment with trypsin caused the cationic fragment to become anionic and abolished the anaphylatoxin but not the chemotactic activity. C3a fragments with identical biologic activity and comparable cationic properties, as determined by acid disc electrophoresis, were obtained by treatment of C3 with C3 convertase, C3 inactivator complex, trypsin, and plasmin. Thrombin produced a similar C3 fragment which was inactive. It was concluded that C3a corresponds to an unusually basic portion of C3 which may be liberated by attack of a variety of enzymes on a highly susceptible region of the native C3 molecule. C3b was cleaved by trypsin and less efficiently by thrombin or plasmin into two antigenically distinct pieces: the larger C3c fragment corresponding to beta(1A) and the smaller C3d fragment to alpha(2D) of aged serum. The c- and the d-fragments were separated and characterized. Isolated C3a rapidly lost its anaphylatoxin activity when treated with small amounts of a partially purified, thermolabile 10S alpha-pseudoglobulin of human serum. The conditions of inactivation suggested an enzymatic reaction. The anaphylatoxin inactivator also destroyed the activity of C5-derived anaphylatoxin and of lysyl bradykinin.
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PMID:Isolation of a fragment (C3a) of the third component of human complement containing anaphylatoxin and chemotactic activity and description of an anaphylatoxin inactivator of human serum. 577 86

Previous studies demonstrated that tissue plasminogen activator-induced fibrinolysis in vitro is retarded in the presence of prothrombin (II) activation and that the anticoagulant-activated protein C appears profibrinolytic by preventing the formation of thrombin (IIa)-like activity during fibrinolysis. To disclose the molecular connection between the generation of IIa and the inhibition of fibrinolysis, a lysis assay that is sensitive to the antifibrinolytic effect of II activation was developed and was used to purify a 60-kDa single-chain protein from human plasma. Because the lysis of a clot, produced from purified components, is retarded when this protein is present and when II activation occurs in situ, the protein was named TAFI (thrombin-activatable fibrinolysis inhibitor). TAFI is cleaved by IIa yielding 35-, 25-, and 14-kDa products. Amino-terminal sequence analyses identified TAFI as a precursor of a plasma carboxypeptidase B (CPB). Formation of the 35-kDa product correlates with both prolongation of lysis time and CPB-like activity. Prolongation of lysis time saturates at about 125 nM TAFI. Activated TAFI inhibits the activation of Glu-plasminogen but does not prolong the lysis of clots formed in the presence of Lys-plasminogen. 2-Guanidinoethylmercaptosuccinic acid, a competitive inhibitor of CPB, completely inhibits prolongation of lysis by activated TAFI in a purified system and the prolongation induced by II activation in barium-adsorbed plasma. This suggests that TAFI accounts for the antifibrinolytic effect that accompanies prothrombin activation and that activated protein C appears profibrinolytic by attenuating TAFI activation.
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PMID:Purification and characterization of TAFI, a thrombin-activable fibrinolysis inhibitor. 778 9

The precursor of plasma carboxypeptidase B (pCPB) also known as thrombin-activable fibrinolysis inhibitor can be converted by thrombin to an active enzyme capable of eliminating C-terminal Lys- and Arg-residues from proteins. The activation is about 1000-fold more efficient in the presence of thrombomodulin (TM). We investigated the antifibrinolytic potency of maximally activated pCPB in plasma and explored the antifibrinolytic mechanism of pCPB. During clotting of plasma in the presence of 3.3 NIH units/ml thrombin and 1 microg/ml soluble TM, more than 80% pro-pCPB was converted into the active form causing an increase of plasma carboxypeptidase activity from 100 units/liter (constitutive activity ascribed to plasma carboxypeptidase N) to 430 units/liter as measured with furoylacroleyl-alanyl-arginine substrate. Under these conditions, lysis of a plasma clot induced by a range of tissue-type plasminogen activator (t-PA) concentrations (0.2-2 microg/ml) was retarded more than 4-fold. A considerable retardation of fibrinolysis was observed upon addition of as little as 12 ng/ml soluble TM, a concentration comparable with physiological concentrations of soluble TM in human plasma. The presence of Ca2+ appeared to be a critical requirement for effective activation of pro-pCPB by thrombin-TM in plasma. Plasminogen-binding sites (C-terminal lysines) on the surface of a plasmin-treated fibrin clot were eliminated within 1-3 min by plasma with maximally activated pCPB, as studied in a recently described model involving fluorescence microscopy. Confocal fluorescence microscopy showed that in the absence of TM plasminogen strongly accumulated on fibrin fibers during t-PA-induced lysis of a plasma clot. In the presence of TM (and a concomitant pro-pCPB activation), lysis was slow and was not accompanied by accumulation of plasminogen on the fibers. In conclusion, generation of active pCPB during clotting of plasma in the presence of Ca2+ and TM leads to a retardation of plasma clot lysis in a wide range of t-PA concentrations, from low to therapeutic, and to a fast elimination of plasminogen-binding sites on partially degraded fibrin. This is a likely mechanism for the antifibrinolytic effect of active pCPB.
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PMID:On the mechanism of the antifibrinolytic activity of plasma carboxypeptidase B. 916 90

Carboxypeptidase U (CPU, EC 3.4.17.20) is a recently described basic carboxypeptidase which circulates in plasma as an enzymatically inactive precursor procarboxypeptidase U (proCPU), also known as plasma carboxypeptidase B precursor or thrombin activatable fibrinolysis inhibitor (TAFI). The activation of the zymogen proceeds through a proteolytic cleavage at Arg-92. The active form - CPU - is able to retard the initial phase of fibrinolysis by cleaving C-terminal lysine residues exposed on fibrin partially degraded by the action of plasmin. These C-terminal lysine residues are essential for the high affinity binding of plasminogen to fibrin and the subsequent activation to plasmin. In this report, the activation of purified human proCPU was studied using trypsin and some key proteases of the coagulation and fibrinolytic cascade, i.e., kallikrein, plasmin and thrombin. The most efficient activation is obtained in the presence of thrombin in complex with thrombomodulin. After in vitro activation, CPU is unstable at 37 degrees C (T(1/2)=15 min). Its stability can be improved dramatically using lower temperatures.
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PMID:Proteolytic activation of purified human procarboxypeptidase U. 1068 74

Carboxypeptidase R (EC 3.4.17.20) (CPR) and carboxypeptidase N (EC 3.4.17.3) (CPN) cleave carboxy-terminal arginine or lysine residues from biologically active peptides such as kinins or anaphylatoxins in the circulation thereby regulating their activities. Although CPN is present in a stable active form in plasma, CPR is generated from proCPR, a plasma zymogen, by proteolytic enzymes such as thrombin, thrombin-thrombomodulin complex and plasmin. We have isolated rat proCPR and CPN cDNA clones which can induce enzymatic activities in culture supernatants of the transfected cells. mRNA of proCPR was detected only in rat liver by Northern hybridization and showed hepatocyte-specific expression. Expression of proCPR mRNA was enhanced following LPS injection, indicating that proCPR production is increased under inflammatory conditions.
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PMID:Molecular cloning and partial characterization of rat procarboxypeptidase R and carboxypeptidase N. 1102 4

Thrombin-activatable fibrinolysis inhibitor (TAFI) is a 60-kDa plasma protein that has been shown to be identical to plasma carboxypeptidase B (CPB) and carboxypeptidase U (CPU). TAFI is activated by thrombomodulin (TM)-bound thrombin and specifically removes the C-terminal Lys and Arg by its CPB activity. One of its target substrates is the C-terminal Lys residue in the alpha-chain of plasmin-digested fibrin, which is critical for plasminogen binding to fibrin. Thus, its removal seems to be the main mechanism through which TAFI inhibits fibrinolysis. In this article, relevance of C-terminal Lys of plasmin-digested fibrin in fibrinolysis is described, and then possible roles of TAFI and TM-bound thrombin in a cross-talk between coagulation and fibrinolysis are discussed.
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PMID:[Regulatory mechanism of fibrinolysis system by thrombin activatable fibrinolysis inhibitor (TAFI)]. 1121 80

When fibrin deposition and removal are properly balanced, the organism is protected from both a catastrophic loss of blood at the site of injury and the inappropriate loss of fluidity within the vascular system. When these activities are not properly balanced, however, severe bleeding or thromboses can occur. Myocardial infarction is a common and morbid consequence of the latter. The thrombin/thrombomodulin complex plays an essential role in regulating this balance because it generates both an anticoagulant substance, activated protein C, and an antifibrinolytic substance, activated TAFI (thrombin activatable fibrinolysis inhibitor, also known as plasma carboxypeptidase B or carboxypeptidase U). Thus, the coagulation and fibrinolytic cascades are explicitly linked by virtue of thrombin catalyzed activation of TAFI, either by the thrombin/thrombomodulin complex or, in the absence of thrombomodulin, by the massive amounts of thrombin generated through the factor XI-dependent pathway after clotting. Some potential targets for diagnosis, prognosis and therapy related to the balance between fibrin formation and removal include: development of a convenient global assay for plasma fibrinolytic potential; an assay for plasma or urine thrombomodulin that had been oxidized at methionine 388 and thereby has lost its capacity to stimulate activation of protein C but not TAFI; an assay for activated TAFI; discovery of a means for tapping the tremendous potential of the vasculature to acutely release tissue-type plasminogen activator; and an assessment of the potential role of polymorphisms in the TAFI gene which might influence TAFI levels or the properties TAFIa. In addition, a much fuller and quantitative understanding of the properties of the coagulation and tibrinolytic cascades is needed in order to optimize diagnosis, prognosis and therapy in disorders such as myocardial infarction that are related to the balance between fibrin formation and removal.
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PMID:Myocardial infarction and the balance between fibrin deposition and removal. 1166 89

Pro-carboxypeptidase R (proCPR), also known as thrombin-activatable fibrinolysis inhibitor (TAFI), precursor of carboxypeptidase U and plasma carboxypeptidase B is present in plasma and following activation by thrombin/thrombomodulin and/or plasmin can remove arginine from the carboxyterminal of C3a and C5a. We have shown that this enzyme can remove terminal arginine from the C5a octapeptide much more efficiently than the classical anaphylatoxin inactivator, carboxypeptidase N (CPN). Since we have previously demonstrated that proCPR is significantly upregulated in the inflammatory state, this enzyme would appear to significantly contribute to the inactivation of C5a, the most potent of the complement derived anaphylatoxins.
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PMID:Inactivation of C3a and C5a octapeptides by carboxypeptidase R and carboxypeptidase N. 1193 78

The Ca(2+)-dependent phospholipid-binding protein annexin II heterotetramer (AIIt) is composed of two copies of annexin II and a p11 dimer. The interaction of the carboxyl-terminal lysine residues of the p11 subunit of AIIt with the lysine-binding kringle domains of plasminogen is believed to play a key role in plasminogen binding and stimulation of the tPA-catalyzed cleavage of plasminogen to plasmin. In the current report, we show that AIIt-stimulated plasminogen activation is regulated by basic carboxypeptidases, in vitro. The incubation of AIIt with a 1/400 molar ratio of carboxypeptidase B for periods as short as 2 min resulted in a significant loss in AIIt-stimulated plasminogen activation. Carboxypeptidase B (CpB) as well as thrombin-activated fibrinolysis inhibitor (TAFIa) and carboxypeptidase N (CpN) rapidly reduced AIIt-stimulated plasminogen activation by 80%. The molar ratio of carboxypeptidase/AIIt for half-maximal inhibition of AIIt was 1/4700, 1/700, and 1/500 for CpB, TAFIa, and CpN, respectively. Treatment of AIIt with carboxypeptidase resulted in loss of both carboxyl-terminal lysine residues from the p11 subunit, which correlated with a decrease in the k(cat) and an increase in the K(m) for plasminogen activation. The data reveal a novel mechanism for the regulation of AIIt-stimulated plasminogen activation.
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PMID:The p11 subunit of annexin II heterotetramer is regulated by basic carboxypeptidase. 1193 91

Carboxypeptidase R (EC 3.4.17.20; CPR) is an unstable basic carboxypeptidase found in fresh serum in addition to carboxypeptidase N (CPN) which is a stable enzyme. CPR in fresh serum is generated from its zymogen (proCPR) during coagulation by trypsin-like enzymes such as thrombin and thrombin/thrombomodulin complexes. Since removal of the C-terminal arginine abrogates the anaphylatoxin activity of C3a and C5a, CPR and CPN are regarded as anaphylatoxin inactivators. We report here that the culture supernatant of activated human neutrophils converts proCPR to CPR. Addition of an elastase specific inhibitor, N-methoxysuccinyl-Ala-Ala-Pro-Val-chloromethyl ketone (MSAAPVCK) to the supernatant of stimulated neutrophils completely inhibited activation of proCPR. On the other hand, a thrombin specific inhibitor, p-Nitrophenyl-p'-amidinophenyl-methanesulfonate hydrochloride (pNP-pAPMS) inhibited only 16% of proCPR activation by the neutrophil supernatant. Furthermore, purified elastase converted proCPR to CPR. Therefore, elastase can activate proCPR directly, or indirectly through activation of some proteases, which have been contaminating in reagents. Release of CPR generating enzymes from neutrophils should play an important role in regulation of excess inflammation.
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PMID:Elastase from activated human neutrophils activates procarboxypeptidase R. 1200 33


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