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:3.4.21.6 (thromboplastin)
13,278 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Based on studies of structure-activity relationship, trans-4-aminomethylcyclohexanecarbonyl-L-phenylalanine-4-carbox ymethylanilide (Tra-Phe-APAA) was designed as a selective plasma kallikrein inhibitor and synthesized. Tra-Phe-APAA inhibited plasma kallikrein with a Ki value of 0.81 microM, while it inhibited glandular kallikrein, plasmin, urokinase, factor Xa and thrombin with Ki values of greater than 500, 390, 200, greater than 500, and greater than 500 microM, respectively. However, its stereoisomer, Tra-D-Phe-APPA did not exhibit any detectable inhibitory activity against the above enzymes.
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PMID:Synthesis of trans-4-aminomethylcyclohexanecarbonyl-L- and -D-phenylalanine-4-carboxymethylanilide and examination of their inhibitory activity against plasma kallikrein. 139 97

A highly selective inhibitor of plasma-kallikrein (PK), N-(trans-4-aminomethylcyclohexylcarbonyl)-L-phenylalanine 4-carboxymethyl-anilide hydrochloride, was designed and synthesized by the authors' group, called PKSI-527 in our laboratories. (I) PKSI-527 inhibited PK with a Ki value of 0.81 microM. By contrast, the Ki values for glandular kallikrein (GK), plasmin, thrombin, urokinase and factor Xa were greater than 500 microM, 390 microM, greater than 500 microM, 200 microM and greater than 500 microM, respectively. (II) Effects of PKSI-527 on bradykinin (BK) generation, coagulation and fibrinolysis by contact activation were examined using human plasma. (a) BK generation induced by kaolin appeared to be reduced by PKSI-527. Furthermore BK generation induced by lambda-carrageenan, a strong inflammatory agent, was also reduced by PKSI-527. (b) Partial thromboplastin time (PTT) was prolonged by PKSI-527, indicating the suppression of the intrinsic coagulation system. (c) Euglobulin clot lysis time (ECLT) of plasma which was shortened by activation with kaolin, was prolonged by the addition of PKSI-527, confirming the participation of PK in contact-fibrinolysis. These results indicate that PKSI-527 shows great potential in elucidating the significance of PK, and as such deserves further investigation.
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PMID:Effect of a highly selective plasma-kallikrein synthetic inhibitor on contact activation relating to kinin generation, coagulation and fibrinolysis. 238 57

The synthetic inhibitors of plasma kallikrein (PK) were found, which are called PKSI-1007, PKSI-0180 and PKSI-0527 in our laboratories. (1) The inhibitors inhibited PK competitively with D-Pro-Phe-Arg-pNA and the Ki values obtained were considerably small, 10(-6) M-10(-7) M. However, the Ki values for glandular kallikrein (GK), plasmin (PL), thrombin (TH) and factor Xa (FXa) were larger. In particular, a selectivity of PKSI-0527 towards PK was very high and the toxicity was weak (i.v. LD50 for mice is over 100 mg/kg). (2) The inhibitors were effective (a) to prevent the bradykinin formation in the kaolin-activated human plasma and the acid-treated ascites taken from the mice bearing Sarcoma 180, (b) to prolong the coagulation time by contact activation, and (c) to inhibit the enhancement of ADP-platelet aggregation by PK. The results indicated that the some PKSI-inhibitors will be much useful for the basic studies, furthermore they deem to be even promising towards the clinical application.
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PMID:Highly selective synthetic inhibitors with regard to plasma-kallikrein activities. 261 76

Inhibitory activities of FUT-187 on trypsin-like serine proteases were compared using camostat mesilate (camostat), and 4-(4-guanidino benzoyloxy)-phenyl acetic acid methanesulfonate (GBPA) known as an active metabolite of camostat in the blood. Ki values of FUT-187 on the competitive inhibition mechanism were 0.097 microM for trypsin, 0.029 microM for pancreatic kallikrein, 0.61 microM for plasma kallikrein, 0.57 microM for plasmin, 2.5 microM for thrombin, 20.4 microM for factor Xa and 6.4 microM for C1r. However, FUT-187 acted as a noncompetitive inhibitor for factor XIIa and an uncompetitive inhibitor for C1s, and Ki values for these proteases were 0.021 and 0.18 microM, respectively. Ki values of camostat for these proteases were in the range of 0.037 to 96.4 microM, and those of GBPA for the above proteases except trypsin and plasma kallikrein were higher than those of FUT-187. The inhibitory activity of FUT-187 on trypsin was not reduced by the addition of the serum at 10%, whereas, that of GBPA was reduced (4.3 fold) in terms of IC50 values. The concentration of FUT-187 required to double APTT (activated partial thromboplastin time) was 1.09 microM, while GBPA, by concentrations up to 1 mM failed to double APTT. The kinin formation by glandular kallikrein in the rat plasma was inhibited by FUT-187 with IC50 value of 0.024 microM, while camostat revealed no inhibition by concentrations up to 1 microM. The complement-mediated hemolyses in the classical and alternative pathways were also inhibited by FUT-187 with IC50 values of 0.17 and 3.5 microM, respectively, the corresponding values for camostat being 350 and 150 microM, respectively. It is concluded that FUT-187 is a potent and selective inhibitor of trypsin-like serine proteases, and its inhibitory activities are stronger than those of camostat on glandular kallikrein, factor XIIa and C1s in complement pathway.
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PMID:[Inhibitory effects of sepimostat mesilate (FUT-187) on the activities of trypsin-like serine proteases in vitro]. 773 78

In a previous report, we described the molecular cloning, expression, and partial characterization of a second human tissue factor pathway inhibitor (TFPI), which we designated as TFPI-2 [Sprecher, C. A., et al. (1994) Proc. Natl. Acad. Sci. U.S.A. 91, 3353-3357]. Recombinant TFPI-2 inhibited the amidolytic activity of trypsin as well as that of factor VIIa in complex with tissue factor. TFPI-2 recently has been shown to be identical to placental protein 5 (PP5), a glycoprotein originally isolated from placenta that exhibits serine protease inhibitory activity. In the present study, we have examined TFPI-2/PP5 for its ability to inhibit a number of serine proteases involved in blood coagulation and fibrinolysis, inasmuch as TFPI-2/PP5 prolonged the coagulation time of human plasma induced by either tissue factor or contact activation in a dose-dependent manner. In addition to its ability to inhibit the amidolytic and proteolytic activities of the factor VIIa-tissue factor complex, TFPI-2/PP5 strongly inhibited the amidolytic activities of human factor XIa, human plasma kallikrein, and human plasmin with Ki values of 15, 25, and 3 nM, respectively. TFPI-2/PP5 was also a weak inhibitor of the activation of factor X by a complex of human factor IXa and poly(lysine) with an apparent Ki of 410 nM. Heparin markedly enhanced the ability of TFPI-2/PP5 to inhibit factor VIIa-tissue factor both in the solution phase and on cell surfaces. In addition, heparin augmented the inhibition of human factor Xa amidolytic activity at relatively high levels (10-100 nM) of TFPI-2/PP5. No significant inhibition of glandular kallikrein, urinary plasminogen activator, tissue plasminogen activator, human activated protein C, human factor Xa, human thrombin, or leukocyte elastase was observed when these proteases were incubated with TFPI-2 in the absence of heparin.
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PMID:Inhibitory properties of a novel human Kunitz-type protease inhibitor homologous to tissue factor pathway inhibitor. 855 84

Ecamulin, a novel prothrombin activating enzyme, has been isolated and purified 63-fold with a 57% yield from the venom of the Middle-Asian sand viper Echis multisquamatus using three-step ion-exchange chromatography. The enzyme was shown to activate prothrombin similarly to Ecarin, a prothrombin-converting enzyme from Echis carinatus venom, however, differing from the latter by structural and physico-chemical properties. The enzyme is a Zn-proteinase: it contains 1 mol Zn per 1 mol of protein. The molecular mass of the enzyme as determined by Sephacryl S-200 chromatography is 93 +/- 2 kDa. Upon SDS-PAAG electrophoresis ecamulin produces two bands with Mr of 67 and 27 kDa under non-reducing conditions, and three bands with Mr of 67, 14 and 13 kDa in the presence of DTT. During native PAGE without SDS, the activator yields one slow mobility band: two bands are observed after addition of DTT or EDTA. Carbohydrates containing N-acetyl-alpha-D-glucosamine residues are localized in the 67 kDa chain. Ecamulin has two isoforms, S2 and S3, that are distinguished by the charge and partial coagulation activities: form S2 has 250 NIH units/mg, while the S3 form has 524 NIH units/mg. The amino acid sequences of the both isoforms are similar but the more active S3 form has 4 times higher content of Gln and 4 times less of Gly than the S2 form. The isoelectric point is 4.3-4.5; E280 of 1% solution is 10.2. Forms S2 and S3 of ecamulin hydrolyze chromogenic substrates of plasma kallikrein S2302 and glandular kallikrein 2266. Ecamulin does not hydrolyze BAEE, TAME, LEE, thrombin substrates Chromozym TH and S2160, factor Xa-S2222, protein Ca-Chromozym PCa and Plasmin S2251. The amidase activity is nonreversibly inhibited by EDTA, o-phenanthroline (the activity is recovered by addition of Zn2+), Cys or DTT, EGTA, DFP, PMSF or pCMB do not inhibit the enzyme activity. Ecamulin converts prothrombin to alpha-thrombin passing by a shunt via the meizothrombin stage. The reaction of prothrombin activation does not require Ca2+, phospholipids of factor Va. Part of this work was presented at the International Conference "Fibrinogen and fibrinolysis", Yalta, September 23-28, 1995.
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PMID:[Isolation and characteristics of ekamulin--a prothrombin activator from multiscaled viper (Echis multisquamatus) venom]. 901 Dec 45

During the course of the development of active center-directed plasmin inhibitors, it was found that N-(trans-4-aminomethylcyclohexanecarbonyl)-lysine-4-methoxycarb onylanilide inhibited plasma kallikrein more potently than other enzymes such as plasmin, urokinase, and thrombin, although the inhibitory activity was not as potent and enzyme selectivity not as high. Based on studies of structure-activity relationship, we designed and synthesized the plasma kallikrein selective inhibitor, N-(trans-4-aminomethylcyclohexanecarbonyl)-phenylalanine-4-carboxy methyl- anilide (Tra-Phe-APAA). Tra-Phe-APAA inhibited plasma kallikrein with a Ki value of 0.81 microM, while it inhibited glandular kallikrein, plasmin, urokinase, tissue plasminogen activator, factor Xa, factor XIIa, and thrombin with Ki values of > 500, 390, 200, > 500, > 500 > 500, and > 500 microM, respectively. We designated Tra-Phe-APAA as PKSI-527. Using PKSI-527 as an affinity ligand, we synthesized a new affinity gel (PKSI-Toyopearl) and employed it for the rapid purification of plasma kallikrein from human plasma. Human plasma activated with kaolin after acid treatment was applied to a PKSI-527-Toyopearl column. Adsorbed protein was eluted with 50 mM glycinehydrochloric acid buffer (pH 3.0). Plasma kallikrein was purified 181-fold with a yield of 85% from the kaolin-activated plasma.
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PMID:Development of plasma kallikrein selective inhibitors. 1038 Mar 51

Human glandular kallikrein 2 (hK2) is a serine protease expressed mainly by the prostate gland with 80% identity in primary structure to prostate specific antigen (PSA). hK2 has proven to be a useful marker of prostate cancer which can be used in combination with PSA to better discriminate between prostate cancer and benign prostate hyperplasia. The studies on hK2 have been hampered by its very low phyciological levels (6 microgram.mL-1), its close similarity to PSA, and the low expression levels obtained using recombinant procedures to produce hK2 (0.7 mg.L-1). We have now generated propeptide mutations of hK2 which can be used to isolate stable, inactive prohK2 mutants. Compared with wild-type hK2, expression of the propeptide hK2 mutants increases the expression levels up to 15-40-fold giving 10-30 mg hK2.L-1. These results indicate that the low expression levels of wild-type hK2 are related to the activation or autoactivation of the wild-type enzyme and the instability of the active protease in cell culture and possibly also in tissue. The purified mutant hK2 may be activated by either enterokinase or factor Xa to generate an enzyme for use in functional studies with the characteristics of the original wild-type protein. Further, the stable inactive mutant hK2 protein may be used for immunizations to generate novel monoclonal antibodies, used as standard material for clinical assays or in crystallization studies where large quantities of protein are required.
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PMID:Production and activation of recombinant hK2 with propeptide mutations resulting in high expression levels. 1058 1

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