Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P00750 (PLA)
 16,800 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Induction of the neutral proteinase, collagenase, is a marker for a specific switch in gene expression observed in rabbit synovial fibroblasts. A variety of agents, including 12-O-tetradecanoylphorbol-13-acetate, cytochalasins B and D, trypsin, chymotrypsin, poly(2-hydroxyethylmethacrylate), and trifluoperazine induced this change in gene expression. Induction of collagenase by these agents was always correlated with a marked alteration in cell morphology, although the cells remained adherent to the culture dishes. The amount of collagenase induced was positively correlated with the degree of shape change produced by a given concentration and, to some extent, with the duration of treatment. Altered cell morphology was required only during the first few hours of treatment with inducing agents; after this time collagenase synthesis continued for up to 6 d even when agents were removed and normal flattened cell morphology was regained. All agents that altered cell morphology also produced a characteristic switch in protein secretion phenotype, characterized by the induction of procollagenase (Mr 53,000 and 57,000) and a neutral metalloproteinase (Mr 51,000), which accounted for approximately 25% and 15% of the protein secreted, respectively. Secretion of another neutral proteinase, plasminogen activator, did not correlate with increased collagenase secretion. In contrast, synthesis and secretion of a number of other polypeptides, including the extracellular matrix proteins, collagen and fibronectin, were concomitantly decreased. That changes in cell shape correlated with a program of gene expression manifested by both degradation and synthesis of extracellular macromolecules may have broad implications in development, repair, and pathologic conditions.
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PMID:Changes in cell shape correlate with collagenase gene expression in rabbit synovial fibroblasts. 632 18

The sequences of urokinase (UK) and tissue-type plasminogen activator (TPA) were aligned with those of chymotrypsin, trypsin, and elastase according to their 'structurally conserved regions'. In spite of its trypsin-like specificity UK was model-built on the basis of the chymotrypsin structure because of a corresponding disulfide pattern. The extra disulfide bond falls to cysteines 50 and 111d. Insertions can easily be accommodated at the surface. As they occur similarly in both, UK and TPA, a role in plasminogen recognition may be possible. Of the functional positions known to be involved in substrate or inhibitor binding, Asp 97, Lys 143 and Arg 217 (Leu in TPA) may contribute to plasminogen activating specificity. PTI binding may in part be impaired by structural differences at the edge of the binding pocket.
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PMID:Adaptation of plasminogen activator sequences to known protease structures. 634 97

The purification and properties of an estradiol-sensitive hydrolytic activity from mouse uterus which fits several criteria for being an induced protein are described. The activity in the uteri of immature animals can be stimulated 2--4-fold by estradiol to that approaching the adult level. Stimulation is blocked by puromycin. The enzyme which we have designated hydrolase II, was purified approx. 400-fold to apparent homogeneity by chromatography on Affigel Blue, DEAE-cellulose and octyl-Sepharose. Hydrolase II is a single chain polypeptide with an estimated mol. wt = 65,000 daltons and has an N-terminal serine residue. A variety of N-blocked L-amino acid nitrophenyl esters are cleaved by the enzyme. Km's at pH 7.2 were all approx. 40 microns. Of substrates tested, phenylalanine nitrophenyl ester had the highest Vmax. Cbz-beta-alanine nitrophenyl ester, which is not a normal protease substrate was cleaved with a Km of 145 microM. The enzyme had no detectable activity against peptide nitroanilide substrates for trypsin-, chymotrypsin- or elastase-like enzymes. It is inhibited by ZPCK and DIFP but not by TLCK and Ala-Ala-Pro-Ala chloromethyl ketone, a potent inhibitor of elastase-like enzymes. Mouse plasma protein protease inhibitors were without effect as was SBTI. Our results rule out hydrolase II being a carnosinase, non-serine esterase, plasminogen activator, collagenase or collagenase activator and suggest that it is a chymotrypsin-like protease.
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PMID:Properties of an estrogen-induced hydrolytic enzyme from mouse uterus. 635 Jul 23

Unlike most proteases, tissue-type plasminogen activator (t-PA) is secreted from cells as an active, single chain "proenzyme" whose catalytic efficiency is comparable with that of the corresponding mature, two-chain enzyme. We have previously suggested that the absence of the "zymogen triad" (Asp194-His40-Ser32; chymotrypsin numbering) contributes to this unusually high enzymatic activity of single chain t-PA. Consistent with this prediction, the single chain form of a variant of t-PA containing the zymogen triad displayed dramatically reduced activity toward synthetic substrates. Activation cleavage of this variant, however, resulted in a mature, two-chain enzyme with full catalytic activity. To further examine the functional significance of the zymogen triad, we used site-specific mutagenesis to construct a variant of t-PA, t-PA/R275E,A292S,F305H, that contained this triad but could not be converted into its two-chain form by plasmin. Characterization of this variant demonstrated that the presence of the zymogen triad specifically suppressed plasminogen activation by single chain t-PA in the absence of fibrin. In addition, these studies indicated that, like wild type t-PA, zymogen activation of this variant could be accomplished by binding to the co-factor fibrin. The combination of full activity in the presence of fibrin and reduced activity in its absence resulted in novel variants of t-PA that displayed dramatically enhanced stimulation by fibrin. While the presence of fibrin increased the catalytic efficiency of t-PA toward plasminogen by a factor of approximately 520, this stimulation factor increased to 130,000 for t-PA/R275E,A292S,F305H. Plasmin-resistant, zymogen-like variants of t-PA, therefore, may represent thrombolytic enzymes with enhanced "clot selectivity."
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PMID:Variants of tissue-type plasminogen activator which display substantially enhanced stimulation by fibrin. 762 53

We investigated the effect of gonadotropins on protease that were suggested to be implicated in the invasive activity of the trophoblast. hCG levels ranging from 10 x 10(3) to 333 x 10(3) IU/L produced a dose-dependent inhibition of the in vitro globinolytic activity of the purified proteases trypsin, chymotrypsin, and urokinase, but failed to inhibit plasmin, collagenase, elastase, and tissue-type plasminogen activator. Likewise, FSH inhibited purified trypsin and urokinase, but not plasmin or tissue-type plasminogen activator. Culture medium conditioned with human trophoblast displayed serine protease and urokinase-like activities; exposure of the cultured trophoblast to exogenous hCG markedly suppressed serine protease and urokinase activities in the conditioned medium. A short treatment of the conditioned medium with trypsin abolished the hCG-mediated inhibition of urokinase activity. The present findings offer an explanation for earlier observations that hCG reduced collagenase activity in trophoblasts without affecting the level of collagenase-specific mRNA. The present results are also consistent with the concept that hCG, by its direct ability to inhibit certain serine proteases and urokinase in trophoblast, suppresses a protease-mediated conversion of procollagenase to active collagenase. The ability of hCG to prevent initiation of the collagenolytic cascade suggests that gonadotropins may regulate the transient invasive activity of the trophoblast.
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PMID:Gonadotropin-mediated inhibition of proteolytic enzymes produced by human trophoblast in culture. 768 89

DX-9065a is an orally active newly synthesized and specific inhibitor for factor Xa. We have examined the property of DX-9065a in vitro and ex vivo. DX-9065a prolonged human plasma recalcification time, APTT and PT. Its doubling concentrations for clotting times of each coagulation assay were 0.49, 0.97 and 0.52 microM, respectively. Kinetic study revealed that DX-9065a inhibited competitively human factor Xa (Ki value: 41 nM). Ki values (microM) for other human serine proteases were as follows; thrombin > 2000, trypsin 0.62, chymotrypsin > 2000, plasmin 23, t-PA 21, plasma kallikrein 2.3 and tissue kallikrein 1000. DX-9065a up to 100 microM had no effects on human platelet aggregation. After intravenous or oral administration, DX-9065a significantly prolonged APTT and PT with a dose dependent manner. These effects were well correlated with anti-Xa activity in plasma. These results suggest that DX-9065a may become an anticoagulant by means of the specific inhibition of factor Xa.
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PMID:DX-9065a, a new synthetic, potent anticoagulant and selective inhibitor for factor Xa. 802 95

Urokinase is a proteinase that normally functions as a plasminogen activator. It is detected in a number of tissues and can be expressed by inflammatory cells such as macrophages and polymorphonuclear leucocytes. Addition of human urokinase to cultures of mucoid or nonmucoid variants of Pseudomonas aeruginosa (strain PAO and clinical isolates from patients with cystic fibrosis) or Pseudomonas cepacia incubated in a minimal medium under nonshaking (oxygen limited) conditions led to dose-dependent enhancement of bacterial growth. The enzyme exhibited a minimal effect on the growth of bacteria when cultured under more intense aeration conditions. This enhancement of bacterial growth by urokinase required the presence of active enzyme and was not detected with inactivated enzyme or noncatalytic domains of the enzyme. Enhancement of bacterial growth was not observed following incubation of P. aeruginosa with other proteinases including thrombin, neutrophil elastase, trypsin, chymotrypsin, or pseudomonas elastase and pseudomonas alkaline proteinase. Therefore, the observed effect of urokinase was relatively specific for this enzyme. As urokinase is a natural constituent of the lung, this enzyme could contribute to bacterial growth during pulmonary infections, particularly in an inflammatory environment in which the oxygen tension may be reduced.
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PMID:Urokinase enhances the growth of Pseudomonas spp. in vitro under nonshaking (oxygen limited) conditions. 803 52

Ecotin, a serine protease inhibitor found in the periplasm of Escherichia coli, has been characterized as an extremely potent anticoagulant and reversible tight-binding inhibitor of human factor Xa (FXa). The ecotin gene was cloned by PCR, highly expressed in E. coli, and purified from the E. coli periplasm. The binding of ecotin to FXa was stoichiometric with an equilibrium dissociation constant Ki of 54 pM. The association rate constant was 1.35 x 10(6) M-1 s-1, and the dissociation rate constant, measured in the presence of human leukocyte elastase (HLE) to prevent reassociation of ecotin with FXa, was 6.5 x 10(-5) s-1. Ecotin prolonged clotting time ca. 10-fold at 0.3 microM and at 2 microM in activated partial thromboplastin time and prothrombin time assays, respectively. Ecotin did not effectively inhibit the human plasma proteases thrombin, tissue factor.factor VIIa, factor XIa, activated protein C, plasmin, or tissue plasminogen activator (t-PA); however, it did potently inhibit factor XIIa, plasma kallikrein, HLE, and bovine trypsin and chymotrypsin. Coincubation of ecotin and FXa at 10 microM each resulted in a (ecotin)2.(FXa)2 complex as determined by gel filtration. Dimerization of ecotin alone was measured by fluorescence titration which yielded a Kd of ca. 390 nM. FXa cleaved ecotin slowly at pH 4.0 between M84 and M85. Replacement of the P1 Met84 residue with Arg and Lys led to FXa inhibitors with Ki values of 11 and 21 pM, respectively. The P1 Arg and Lys mutants also significantly inhibited thrombin, factor XIa, activated protein C, plasmin, factor XIIa, kallikrein, and bovine trypsin and chymotrypsin but did not inhibit tissue factor.factor VIIa, t-PA, or HLE.
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PMID:Ecotin is a potent anticoagulant and reversible tight-binding inhibitor of factor Xa. 814 99

Unlike most serine proteases of the chymotrypsin family, tissue-type plasminogen activator (tPA) is secreted from cells as an active, single-chain enzyme with a catalytic efficiency only slightly lower than that of the proteolytically cleaved form. A zymogenic mutant of tPA has been engineered that displays a reduction in catalytic efficiency by a factor of 141 in the single-chain form while retaining full activity in the cleaved form. The residues introduced in the mutant, serine 292 and histidine 305, are proposed to form a hydrogen-bonded network with aspartate 477, similar to the aspartate 194-histidine 40-serine 32 network found to stabilize the zymogen chymotrypsinogen.
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PMID:Converting tissue plasminogen activator to a zymogen: a regulatory triad of Asp-His-Ser. 821 Nov 62

Six monoclonal IgG1-k antibodies (LK2, LK3r, LK4-55, LK5, LK6-55, LK7r) were raised against platelet membrane GPIIIa in order to study the structure-function relationship of this molecule. Antibodies were selected on their ability to react with GPIIIa by ELISA on adherent platelets, by immunoblot on platelet lysates and by fluorescence flow cytometry on intact platelets. Fluorescence reactivity varied from 3- to 202-fold greater than isotype control fluorescence. Two MoAbs reacted on immunoblot under reduced conditions (LK7r and LK3r). Two reacted with a 55 kD chymotrypsin/subtilisin digest of GPIIIa which is likely to exclude amino acids 121-348 (LK4-55 and LK6-55). Four of the MoAbs (LK5, LK3r, LK2 and LK4-55) inhibited tyrosine phosphorylation of one to four distinct bands on immunoblot. LK4-55 reacted with an N-terminal 66 amino acid fusion protein of GPIIIa near the PLA epitope (Leu 33). LK7r reacted with a 212-222 peptide reported to be an RGD fibrinogen binding site. LK2 reacted near a disintegrin-RGD binding site. Except for LK5, all inhibited ADP, collagen and thrombin-induced platelet aggregation in a heterogeneous fashion. Percentage inhibition of 125I-fibrinogen binding to platelets varied from 18% to 98%. No correlation was noted between inhibition of fibrinogen binding, location of MoAb binding on GPIIIa, reactivity of MoAb binding with GPIIIa, inhibition of thrombin-induced tyrosine phosphorylation or inhibition of platelet aggregation induced by ADP, collagen or thrombin. Thus MoAbs, binding to platelet GPIIIa at different sites, inhibit platelet aggregation in a heterogeneous manner.
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PMID:Heterogenous inhibition of platelet aggregation by monoclonal antibodies binding to multiple sites on GPIIIa. 854 51


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