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:3.5.1.4 (deaminase)
5,113 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

An anticoagulant isolated from salivary gland extracts of the ixodid tick Rhipicephalus appendiculatus was purified by gel filtration on Sephadex G-100, ion exchange on DEAE-cellulose, aprotinin-Sepharose, and by high-pressure-liquid size-exclusion chromatography. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that the anticoagulant activity was associated with a protein of an apparent Mr of 65 kDa. The purified molecule had a pI in the range of 8.0-8.5 on chromatofocusing and was stable over a wide pH range, but was heat labile and susceptible to inactivation by trypsin and reductive alkylation. The anticoagulant did not inhibit thrombin-initiated fibrin formation and had no detectable fibrino(geno)lytic or phospholipase-like activities. Although it inhibited factor Xa-induced clotting of bovine plasma, it did not affect the amidase activity of factor Xa toward a synthetic substrate, suggesting that the anticoagulant acts at a site distinct from the active site of factor Xa or on other components of the prothrombinase complex.
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PMID:Purification and characterization of an anticoagulant from the salivary glands of the ixodid tick Rhipicephalus appendiculatus. 202 16

The amidase activity of human alpha-thrombin has been studied in the presence of the adenosine nucleotides AMP, ADP and ATP. At low concentrations, adenosine nucleotides increase thrombin activity up to 30%, while at high concentrations (greater than 5 mM) inhibition takes place up to 20%. Inhibition is progressively reduced by increasing substrate concentration. A simple, phenomenological description of the linkage between adenosine nucleotide binding and amidase activity of human alpha-thrombin is proposed and the free energy changes for the underlying reactions involved in the linkage scheme are resolved by global analysis of the experimental data. The linkage scheme assumes that thrombin activation is determined by a conformational transition due to binding of adenosine nucleotides to a regulatory site. Inhibition, on the other hand, would be a consequence of competitive binding to the catalytic site.
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PMID:The linkage between adenosine nucleotide binding and amidase activity in human alpha-thrombin. 220 77

The amidase activity of human alpha-thrombin has been studied in the pH range 5.5 to 10, and at four different chloride concentrations from 5 mM to 1 M. The Michaelis-Menten constant, Km, shows a bell-shaped dependence over the pH range studied, with a minimum around pH 8. The pH dependence of the catalytic constant, kcat, shows multiple inflection points especially at low (less than 0.1 M) chloride concentrations, thereby implicating the existence of multiple catalytic forms of the enzyme. A general linkage scheme is proposed for the analysis of the effect of protons on thrombin amidase activity, and experimental data have globally been analysed over the entire pH range in terms of such a scheme. Four proton-linked ionizable groups seem to be involved in the control of thrombin amidase activity. Two of these groups change their pK value upon substrate binding to the enzyme and account for the pH dependence of Km. All four groups control the catalytic activity of the enzyme which decreases with increasing protonation. Chloride has little effect on Km, while kcat changes significantly at pH less than 8. This effect is due to an increased enzymatic activity of the highly protonated intermediates at high chloride concentrations, as well as to the pK shift of two proton-linked ionizable groups.
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PMID:Effect of protons on the amidase activity of human alpha-thrombin. Analysis in terms of a general linkage scheme. 226 57

The A- and B-chains have been isolated from the non-covalent complex of human thrombin A- and B-chains, using selective reduction of the interchain disulfide bridge. The B-chain thus isolated (de-A-thrombin) retains its conformation, which is close to the native one and thus differs considerably from the B-chain isolated from the fully reduced enzyme. Nevertheless, the proteolytic (in terms of fibrinogen clotting) and amidase activities of de-A-thrombin are markedly reduced as compared to the native enzyme and the non-covalent complex of A- and B-chains. It is assumed that the A-chain of thrombin is necessary for normal functioning of the active site of thrombin localized in the B-chain.
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PMID:[Enzymatic properties of the isolated B-chain of human thrombin]. 275 67

gamma-Thrombin was produced during autolysis or limited proteolysis of coagulant gamma-thrombin. This thrombin form loses its ability to coagulate fibrinogen but preserves the esterase and amidase activity on the low-molecular-weight synthetic substrates. This evidences for the integrity of the active site of gamma-thrombin and for the integrity break of the enzyme molecule region responsible for the binding with fibrinogen. gamma-Thrombin with the minimal coagulating activity, possessing high esterase and amidase activity is obtained. Fibrin-agarose possessing affinity to gamma-thrombin and specifically not binding gamma-thrombin was used to remove admixtures of the coagulant gamma-thrombin from the preparations of gamma-thrombin obtained during the enzyme autolysis.
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PMID:[Affinity chromatography of human gamma-thrombin on fibrin-agarose]. 318 51

The toxic and biological activities of four samples of Trimeresurus purpureomaculatus venom were examined. The lethality, protein composition and biological activities of the four venom samples were similar. Three of the venom samples had LD50 (i.v.) values of 0.9 micrograms/g while the fourth had a lower LD50 (i.v.) of 0.45 micrograms/g. All four venom samples exhibited hemorrhagic, edema-inducing, anticoagulant and thrombin-like activities as well as the usual enzymes found in crotalid venoms. DEAE-Sephacel ion exchange chromatographic fractionation of the venom yielded 10 protein fractions. Only two fractions (fractions A and F) were lethal to mice; the major lethal fraction being fraction F. This fraction had an LD50 (i.v.) of 0.2 micrograms/g and exhibited hemorrhagic, edema-inducing and thrombin-like activity. It also exhibited phospholipase A, arginine ester hydrolase, arginine amidase, protease, 5'-nucleotidase, acetylcholinesterase and alkaline phosphomonoesterase activities. The lethal potency of fraction F is potentiated by fraction G, which exhibited anticoagulant activity as well as hemorrhagic, edema-inducing and enzymatic activities. Fractions F plus G account for almost 100% of the lethal potency of the venom.
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PMID:Biological properties of Trimeresurus purpureomaculatus (shore pit viper) venom and its fractions. 324 58

A low molecular weight protein protease inhibitor was purified from Japanese horseshoe crab (Tachypleus tridentatus) hemocytes. It consisted of a single polypeptide with a total of 61 amino acid residues. This protease inhibitor inhibited stoichiometrically the amidase activity of trypsin (Ki = 4.60 X 10(-10) M), and also had inhibitory effects on alpha-chymotrypsin (Ki = 5.54 X 10(-9) M), elastase (Ki = 7.20 X 10(-8) M), plasmin, and plasma kallikrein. However, it had no effect on T. tridentatus clotting enzyme and factor C, mammalian blood coagulation factors (activated protein C, factor Xa and alpha-thrombin), papain, and thermolysin. The complete amino acid sequence of this inhibitor was determined and its sequence was compared with those of bovine pancreatic trypsin inhibitor (BPTI) and other Kunitz-type inhibitors. It was found that the amino acid sequence of this inhibitor has a high homology of 47 and 43% with those of sea anemone inhibitor 5-II and BPTI, respectively. Thus, this protease inhibitor appeared to be one of the typical Kunitz-type protease inhibitors.
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PMID:Purification and amino acid sequence of Kunitz-type protease inhibitor found in the hemocytes of horseshoe crab (Tachypleus tridentatus). 330 64

Bacterial protein, staphylocoagulase, binds stoichiometrically to human prothrombin resulting in a coagulant complex, staphylothrombin. The enzymatic properties of staphylothrombin differ from those of alpha-thrombin in their substrate specificities toward natural and synthetic substrates, in addition to their interaction with protease inhibitors. In order to obtain information about the region of staphylocoagulase that interacts with human prothrombin, staphylocoagulase was cleaved by alpha-chymotrypsin. This limited alpha-chymotryptic cleavage of staphylocoagulase yielded three large fragments, fragments of 43, 30, and 20 kDa. The 43-kDa fragment exhibited a high affinity for human prothrombin (Kd = 1.7 nM), which is comparable to the affinity observed using intact staphylocoagulase (Kd = 0.46 nM). A complex of the 43-kDa fragment and prothrombin possessed both clotting and amidase activities essentially identical to those observed in a complex of intact staphylocoagulase and prothrombin. The 30-kDa fragment exhibited weaker affinity for prothrombin (Kd = 120 nM). While a complex of this fragment and prothrombin did not exhibit clotting activity, it nonetheless possessed a weak amidase activity. The 20-kDa fragment was found only to bind to prothrombin. The NH2-terminal sequence analyses of these fragments revealed that the 43-kDa fragment constitutes the NH2-terminal portion of staphylocoagulase, and contains the 30-kDa and 20-kDa fragments. It is therefore concluded that the functional region of staphylocoagulase for binding and activation of prothrombin is localized in the NH2-terminal region of the intact protein. The 43-kDa fragment contains 324 amino acids with a molecular weight of 38,098. The 43-kDa fragment has an unusual amino acid composition based on the sequence, in which the sum of Asp (28 residues), Asn (22), Glu (35), Gln (9), and Lys (52) residues accounts for more than 45% of the total residues. A comparison of the amino acid sequence of the 43-kDa fragment with that of streptokinase did not reveal any obvious sequence homology. There was also no sequence homology with those of trypsin, alpha-chymotrypsin, and elastase.
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PMID:Difference in enzymatic properties between "staphylothrombin" and free alpha-thrombin. 355 30

A thrombin-like enzyme was isolated in 6% yield from the venom of Crotalus durissus terrificus by ammonium sulfate precipitation followed by gel filtration on Sephadex G-75 and finally affinity chromatography on Sepharose-1,4-butanediol-diglycyl-p-aminobenzamide eluted with 0.15 M benzamidine. The enzyme behaved like a single component on SDS-PAGE corresponding to a molecular weight of 34 kDa. The specific activity of the enzyme toward bovine fibrinogen was 71 NIH U/mg protein. The pH optimum for the coagulation of human fibrinogen was 8.0. The enzyme hydrolyzes the alpha-chain of fibrinogen, has amidase activity on L-arginine-p-nitroanilide and L-arginine-7-amido-4-methyl-coumarin amino terminal blocked peptides and presents esterolytic activity on N-alpha-tosyl-L-arginine-methylester.
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PMID:Isolation and characterization of a thrombin-like enzyme from the venom of Crotalus durissus terrificus. 359

This report documents attempts to mimic the rate enhancement effect of thrombomodulin on human alpha-thrombin-catalyzed activation of human protein C in the absence of exogenous calcium. Specifically the following tryptamine analogs at 1 mM concentration were shown to enhance the protein C activation rate relative to a control with no added effector at pH 8.3 (50 mM Tris-HCl, 0.1 M NaCl, 37 degrees C): serotonin, 1.2; tryptamine, 2.9; 5-fluorotryptamine, 4.4; 6-fluorotryptamine, 7.2. At much higher levels, e.g. 10 mM, all of the above effectors, as well as indole, showed a moderate inhibition of human protein C activation. ATP, a platelet release product, showed a sigmoidal inhibition pattern similar to that found previously for thrombin amidase, clotting, and esterase activity (Conery, B.G., and Berliner, L.J. (1983) Biochemistry 22, 369-375). Overall, the enhancement factors for human alpha-thrombin activation of protein C with the tryptamine analogs described above were remarkable when considering the effect of a simple ligand versus the natural activator, thrombomodulin.
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PMID:Ligands which effect human protein C activation by thrombin. 365 43


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