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:4.2.2.7 (heparinase)
1,270 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The genome of hepatitis C virus (HCV) encodes two envelope glycoproteins (E1 and E2), which are thought to be responsible for receptor binding and membrane fusion resulting in virus penetration. To investigate cell surface determinants important for HCV infection, we used a recombinant vesicular stomatitis virus (VSV) in which the glycoprotein gene was replaced with a reporter gene encoding green fluorescent protein (GFP) and produced HCV-VSV pseudotypes possessing chimeric HCV E1 or E2 glycoproteins, either individually or together. The infectivity of the pseudotypes was determined by quantifying the number of cells expressing the GFP reporter gene. Pseudotypes that contained both of the chimeric E1 and E2 proteins exhibited 10--20 times higher infectivity on HepG2 cells than the viruses possessing either of the glycoproteins individually. These results indicated that both E1 and E2 envelope proteins are required for maximal infection by HCV. The infectivity of the pseudotype virus was not neutralized by anti-VSV polyclonal antibodies. Bovine lactoferrin specifically inhibited the infection of the pseudotype virus. Treatment of HepG2 cells with Pronase, heparinase, and heparitinase but not with phospholipase C and sodium periodate reduced the infectivity. Therefore, cell surface proteins and some glycosaminoglycans play an important role in binding or entry of HCV into susceptible cells. The pseudotype VSV possessing the chimeric HCV glycoproteins might offer an efficient tool for future research on cellular receptors for HCV and for the development of prophylactics and therapeutics for hepatitis C.
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PMID:Characterization of pseudotype VSV possessing HCV envelope proteins. 1148 95

Antithrombin inhibits chemokine-induced migration of neutrophils by activating heparan sulfate proteoglycan-dependent signaling. Mechanisms of antithrombin's effects on neutrophils were, therefore, studied by testing function and expression of heparan sulfate proteoglycans in RT-PCR or flow cytometry and cell migration assays, respectively. In vitro effects of antithrombin on human neutrophil migration in modified Boyden chambers were abolished by pretreating cells with heparinase-1, chondroitinase, sodium chlorate, and anti-syndecan-4 antibodies. Expression of syndecan-4 mRNA and protein in neutrophils was demonstrated in RT-PCR and anti-syndecan-4 immunoreactivity assay, respectively. In the presence of pentasaccharide, antithrombin lost its activity on the cells. Data suggest that antithrombin regulates neutrophil migration via effects of its heparin-binding site on cell surface syndecan-4.
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PMID:Syndecan-4 as antithrombin receptor of human neutrophils. 1154 50

Antithrombin inhibits chemokine-induced migration of neutrophils by activating heparan sulfate proteoglycan-dependent signaling. Whether antithrombin affects migration of other types of leukocytes is not known. We investigated the effects of antithrombin on spontaneous and chemokine-triggered migration of lymphocytes and monocytes from human peripheral blood in modified Boyden chamber micropore filter assays. Lymphocyte and monocyte populations from human peripheral blood were purified using magnetic antibody cell sorting. The signaling mechanisms required for antithrombin-dependent migration were studied using signaling enzyme blockers. Expression of heparan sulfate proteoglycan core protein was studied by RT-PCR and flow cytometry. The antithrombins used were Kybernin P from human plasma and a monoclonal-antibody-purified preparation from this plasma. Pretreatment of lymphocytes and monocytes with antithrombin inhibited chemotaxis toward optimal concentrations of interleukin-8 or Rantes (regulated upon activation normal T-cell expressed and activated) at concentrations of antithrombin as low as 10 nU/ml. In the absence of the chemokines, direct exposure of cells to gradients of antithrombin stimulated migration. Effects of antithrombin were abolished by pretreating cells with heparinase-1, chondroitinase, sodium chlorate and anti-syndecan-4 antibodies. Expression of syndecan-4 mRNA and protein in monocytes and lymphocytes was demonstrated in RT-PCR and anti-syndecan-4 immunoreactivity assays, respectively. In the presence of pentasaccharide, antithrombin lost its effect on cells. Data indicate that antithrombin directly inhibits chemokine-stimulated migration of monocytes and lymphocytes via the effects of its heparin-binding site on cell surface syndecan-4 by activation of protein kinase C and Rho signaling.
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PMID:Syndecan-4 mediates antithrombin-induced chemotaxis of human peripheral blood lymphocytes and monocytes. 1180 40

The porcine reproductive and respiratory syndrome virus (PRRSV) has a very restricted tropism for well-differentiated cells of the monocyte-macrophage lineage, which is probably determined by specific receptors on these cells. In this study, the importance of heparinlike molecules on porcine alveolar macrophages (PAM) for PRRSV infection was determined. Heparin interacted with the virus and reduced infection of PAM up to 92 or 88% for the American and European types of PRRSV, respectively. Other glycosaminoglycans, similar to heparin, had no significant effect on infection while heparinase treatment of PAM resulted in a significant reduction of the infection. Analysis of infection kinetics showed that PRRSV attachment to heparan sulfate occurs early in infection. A heparin-sensitive binding step was observed which converted completely into a heparin-resistant binding after 120 min at 4 degrees C. Using heparin-affinity chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), it was observed that the structural matrix (M) and nucleocapsid (N) proteins attached to heparin. Nonreducing SDS-PAGE revealed that M bound to heparin mainly as a complex with glycoprotein GP(5) and that the N protein bound to heparin as a homodimer. GP(3), which was identified as a minor structural protein of European types of PRRSV, did not bind to heparin. Since the N protein is not exposed on the virion surface, it was concluded that the structural M protein and the M-GP(5) complex contribute to PRRSV attachment on a heparinlike receptor on PAM. This is the first report that identifies a PRRSV ligand for a cell surface heparinlike receptor on PAM.
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PMID:Involvement of the matrix protein in attachment of porcine reproductive and respiratory syndrome virus to a heparinlike receptor on porcine alveolar macrophages. 1193 97

Non-infectious, envelope protein-free, retrovirus-like particles (VLP) derived from either Moloney murine leukemia virus (MLV) or human HIV are able to bind efficiently to, but not infect, target cells. Upon subsequent addition to the bound particles of the G protein of vesicular stomatitis virus (VSV-G), an efficient surrogate retrovirus envelope protein, the VLP are efficiently taken up by the cells to produce infection. Cell attachment of the VLP is efficiently inhibited by soluble heparin and dextran sulfate and less efficiently abrogated by several other glycosaminoglycans (GAGs) including chondroitin sulfate A and chondroitin sulfate B (dermatan sulfate), as determined by deconvolution microscopic immunodetection of the viral gag protein and by quantitative binding studies of metabolically labeled (35)S-VLP. Enzymatic digestion of heparan sulfate (HS) from the cell surface with heparinase I also reduces VLP binding. Furthermore, VLP adsorption onto several CHO cell lines variably deficient in cell surface GAG is significantly but incompletely abrogated. De-sulfated heparins are less efficient than native heparin in inhibiting the Polybrene-mediated binding of VLP, whereas growth of human cells in the presence of sodium chlorate leads to significant reduction of Polybrene-mediated VLP binding. In addition, specific inhibition of VLP binding and infectivity of mature infectious VSV-G-pseudotyped virus is observed in the presence of heparin and HS under Polybrene-free conditions. We conclude from these studies that the presence of Polybrene, the degree of sulfation of cell surface GAG, and possibly the presence of charged cell surface macromolecules create an electrostatic environment that promotes optimum binding of VLP to cells. Additionally, our results demonstrate that, in the absence of Polybrene, initial attachments of non-infectious, envelope protein-free VLP and probably mature infectious virus particles are mediated by interactions of the virus particles with cell surface heparan sulfate, and possibly with other GAG molecules.
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PMID:Cell surface heparan sulfate is a receptor for attachment of envelope protein-free retrovirus-like particles and VSV-G pseudotyped MLV-derived retrovirus vectors to target cells. 1199 44

Sulfo-glycosaminoglycans (sGAGs) are involved in the assembly of tau in at least a subpopulation of paired helical filaments (PHFs) in Alzheimer's disease (AD). To further understand the role of sGAG molecules in the structure of PHFs, we isolated PHFs from patients with AD and treated them with heparinase. Immunoelectron microscopy and Western blotting (WB) were used later on to analyze the changes obtained. The heparinase treatment abolished Tau14 and AT8 immunodecoration (two N-terminal tau antibodies) and increased PHF-1 labeling (a C-terminal antibody). In addition, heparinase-treated filaments are more labile than control ones as demonstrated by sodium dodecyl sulfate-extraction and subsequent WB. In summary, our results demonstrate that sGAG content affects PHF conformation as well as PHF-tau solubilization.
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PMID:Sulfo-glycosaminoglycan content affects PHF-tau solubility and allows the identification of different types of PHFs. 1206 74

Our current studies have demonstrated that human parainfluenza virus type 3 (HPIV-3) utilizes heparan sulfate (HS) for its efficient cellular entry. HPIV-3 interacted with HS-agarose in vitro and the cellular entry and infection of HPIV-3 were reduced following (a) infection of human epithelial lung A549 cells with HPIV-3 pre-incubated with soluble HS; (b) treatment of A549 cells with heparinase to remove cell surface HS and sodium chlorate (NaClO(3)), a potent inhibitor of proteoglycan sulfation; and (c) infection of HS-deficient mutant CHO cell lines. However, in each instance, complete inhibition of HPIV-3 entry did not occur, suggesting the presence of additional nonproteoglycan cell surface molecule(s) that is required for HPIV-3 entry. Thus the cell surface HS appears to play an important role in efficient cellular entry of HPIV-3.
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PMID:Role of heparan sulfate in human parainfluenza virus type 3 infection. 1209 75

Heparin and heparan sulfate fragments, obtained by bacterial heparinase and heparitinases, bearing an unsaturation at C4-C5 of the uronic acid moiety, are able to produce up to 80% reduction of the cytosolic calcium of smooth muscle cell lines. Unsaturated disaccharides from chondroitin sulfate, dermatan sulfate, and hyaluronic acid are inactive, indicating that, besides the unsaturation of the uronic acid, a vicinal 1 --> 4 glycosidic linkage is needed. An inverse correlation between the molecular weight and activity is observed. Thus, the ED(50) of the N-acetylated disaccharide derived from heparan sulfate (430 Da) is 88 microm compared with 250 microm of the trisulfated disaccharide (650 Da) derived from heparin. Except for enoxaparin (which contains an unsaturation at the non-reducing end and 1 --> 4 glycosidic linkage), other low molecular weight heparins and native heparin are practically inactive in reducing the cytosolic calcium levels. Thapsigargin (sarcoplasmic reticulum Ca(2+)-ATPase inhibitor), vanadate (cytoplasmic membrane Ca(2+)-ATPase inhibitor), and nifedipine and verapamil (Ca(2+) channel antagonists) do not interfere with the effect of the trisulfated disaccharide upon the decrease of the intracellular calcium. A significant decrease of the activity of the trisulfated disaccharide is observed by reducing extracellular sodium, suggesting that the fragments might act upon the Na(+)/Ca(2+) exchanger promoting the extrusion of Ca(2+). This was further substantiated by binding experiments and circular dichroism analysis with the exchanger inhibitor peptide.
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PMID:Heparin and heparan sulfate disaccharides bind to the exchanger inhibitor peptide region of Na+/Ca2+ exchanger and reduce the cytosolic calcium of smooth muscle cell lines. Requirement of C4-C5 unsaturation and 1--> 4 glycosidic linkage for activity. 1237 9

The cellular mechanism by which apolipoprotein E (apoE) affects the pathogenesis of Alzheimer's disease (AD) is not understood. We have examined the effect of apolipoprotein E on the internalization of exogenous amyloid-beta 1-40 (Abeta40) into a rat brain crude synaptosomal preparation. Abeta40 peptide in soluble (within 1 h of dilution in buffer) or aggregated (aged 4 days before dilution in buffer) form was pre-incubated with lipidated apoE then added to synaptosomes; intraterminal amyloid-beta labeling was quantified using flow cytometry following immunolabeling with the anti-Abeta (10G4) antibody. The number of Abeta-positive synaptosomes was increased ( approximately 50%) by treatment with a soluble Abeta/apoE mixture compared with treatment with soluble Abeta40 alone. However, when the Abeta was aggregated, less sodium dodecyl sulfate (SDS)-stable Abeta/apoE complex was formed and the addition of apoE decreased the number of Abeta-positive terminals. The addition of the lipoprotein-receptor related protein (LRP) antagonist receptor-associated protein (RAP) inhibited the apoE-induced increase in synaptosomal Abeta, and controls treated with trypsin and heparinase confirm intraterminal localization of the majority of the soluble Abeta. The apoE-mediated increase in Abeta labeling was confirmed in intact cells by immunocytochemistry of dorsal root ganglion (DRG) neurons. These results suggest that complex formation with apoE enhances internalization of soluble Abeta uptake into terminals.
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PMID:Apolipoprotein E enhances uptake of soluble but not aggregated amyloid-beta protein into synaptic terminals. 1261 44

During prion diseases, the host protein PrPC is refolded into an abnormal conformer "prion" PrPSc. Histological and pharmacological data have suggested that glycosaminoglycans may be involved in the development of prion diseases. Here we present the first direct evidence that cellular glycosaminoglycans play a role in the biogenesis of PrPSc in prion-infected ScN2a cells. When ScN2a cells were incubated with estradiol beta-d-xyloside to inhibit the glycosylation of proteoglycans, PrPSc was vastly reduced. Treating ScN2a-M cells with heparinase III, but not with heparinase I or chondroitinase ABC, caused a profound reduction of PrPSc. In contrast, neither the amount of PrPC nor its subcellular distribution were affected as assayed by immunofluorescence microscopy and flotation procedures. In vitro treatment of ScN2a membranes with heparinase III at either neutral or acidic pH did not reduce the level of protease-resistant PrPSc. The inhibitor of sulfation, sodium chlorate, vastly reduces PrPSc in ScN2a cells (Gabizon, R., Meiner, Z., Halimi, M., and Ben-Sasson, S. A. (1993) J. Cell. Physiol. 157, 319-325). Both soluble heparan sulfate and chondroitin sulfate partially restored the level of PrPSc in chlorate-treated cells. We conclude that heparinase III-sensitive, presumably undersulfated, cellular heparan sulfate plays a significant role in the biogenesis of PrPSc in ScN2a cells.
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PMID:Cellular heparan sulfate participates in the metabolism of prions. 1287 49


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