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 binding of Apolipoprotein E supplemented triglyceride emulsions to sulfated glycosaminoglycans demonstrated specificity for the carbohydrate polymers. Glucosamine containing glycosaminoglycans with relatively less sulfate had little affinity for the Apo E emulsion whereas those with more sulfate (i.e. heparin and sulfated heparans) effectively bound the emulsion. Galactosamine containing glycosaminoglycans (chondroitin 4 sulfate and dermatan sulfate) demonstrated no binding. The Apo E induced uptake of triglyceride emulsions by hepatocytes was inhibited by highly sulfated polysaccharides (i.e. heparin, dextran sulfate) but other glycosaminoglycans which did not bind the emulsion were ineffective in this inhibition. The same sulfated compounds which inhibited the hepatocyte Apo E emulsion interaction effectively released hepatic lipase from isolated heptic perfusions. Glycosaminoglycan sulfates which did not bind the Apo E supplemented emulsions and did not inhibit hepatocyte association were ineffective in releasing lipase. A heparan mixture isolated from human liver was much less effective in inhibiting Apo E induced association of emulsions with hepatocytes, than heparin. A highly sulfated octasaccharide fraction isolated from bovine liver heparin inhibited more effectively than the human heparans but less than the heparin. Inhibition of Apo E mediated hepatocyte emulsion association was produced by a one hour exposure of the cells to either heparinase or heparanase. The heparanase was more active than the heparinase and both were effective in the presence of protease inhibitors. Enzymes hydrolyzing chondroitin sulfates and hyaluronic acid were ineffective in inhibiting the Apo E induced association. The specific binding of human low density lipoprotein to the hepatocyte was much less effected by the heparanase exposure than the Apo E mediated binding.
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PMID:The relevance of glycosaminoglycan sulfates to Apo E induced lipid uptake by hepatocyte monolayers. 294 1

The effects of rabbit T lymphocytes on rabbit aortic smooth muscle cell (SMC) phenotype and proliferation were investigated in vitro. SMCs seeded at confluent density in primary culture had a volume fraction of myofilaments (Vvmyo) of 49.8 +/- 2.6% after 3 days of culture, not significantly different from that of freshly dispersed cells (Vvmyo, 54.1 +/- 2.1%). Sister cultures of SMCs to which Concanavalin A-activated T lymphocytes or T lymphocyte-conditioned medium was added had significantly lower Vvmyo (35.5 +/- 2.2% and 31.6 +/- 2.3%, respectively) at the same time point. We have previously shown that a decrease in Vvmyo could be induced by the heparan sulfate-degrading activity of living macrophages and by commercial preparations of heparinase. While activated T lymphocytes also completely degraded heparan sulfate-rich 35S-labeled extracellular matrix (an effect inhibited by the addition of 10 micrograms/mL heparin), no heparanase-like activity was detected in T lymphocyte-conditioned medium, indicating that for this cell type SMC phenotypic change is induced by a different mechanism. Incubation of the T lymphocyte-derived cytokine interferon gamma (IFN-gamma) with freshly isolated rat SMCs caused a significant reduction in Vvmyo at day 2 in primary culture from 54.3 +/- 2.1% (control) to 35.4 +/- 3.0%. Furthermore, a neutralizing antibody specific for IFN-gamma removed the effect of T lymphocytes and medium conditioned by them, thus positively identifying IFN-gamma as the T lymphocyte factor responsible for this activity. T lymphocyte-conditioned medium was mitogenic for passaged (low Vvmyo) SMCs.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:T lymphocytes affect smooth muscle cell phenotype and proliferation. 762 15

The cause and consequence of altered proteoglycans in atherosclerosis are poorly understood. To determine whether proteoglycans affect monocyte binding, we studied the effects of heparin and proteoglycan degrading enzymes on THP-1 monocyte adhesion to subendothelial matrix (SEM). Monocyte binding increased about 2-fold after SEM was treated with heparinase. In addition, heparin decreased monocyte binding to fibronectin, a known SEM protein, by 60%. These data suggest that SEM heparan sulfate inhibits monocyte binding to SEM proteins. We next examined whether lysolecithin, a constituent of modified lipoproteins, affects endothelial heparan sulfate proteoglycan (HSPG) production and monocyte binding. Lysolecithin (10-200 microM) decreased total 35SO4 in SEM (20-75%). 2-fold more monocytes bound to SEM from lysolecithin treated cells than to control SEM. Heparinase treatment did not further increase monocyte binding to lysolecithin-treated SEM. HSPG degrading activity was found in medium from lysolecithin-treated but not control cells. 35SO4-labeled products obtained from labeled matrix treated with lysolecithin-conditioned medium were similar in size to those generated by heparinase. These data suggest that lysolecithin-treated endothelial cells secrete a heparanase-like activity. We hypothesize that decreased vessel wall HSPG, as occurs in atherogenic conditions, allows increased monocyte retention within the vessel and is due to the actions of an endothelial heparanase.
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PMID:Lysolecithin-induced alteration of subendothelial heparan sulfate proteoglycans increases monocyte binding to matrix. 853 Mar 67

Vessel wall subendothelial extracellular matrix, a dense mesh formed of collagens, fibronectin, laminin, and proteoglycans, has important roles in lipid and lipoprotein retention and cell adhesion. In atherosclerosis, vessel wall heparan sulfate proteoglycans (HSPG) are decreased and we therefore tested whether selective loss of HSPG affects lipoprotein retention. A matrix synthesized by aortic endothelial cells and a commercially available matrix (Matrigel; , Rutherford, NJ) were used. Treatment of matrix with heparinase/heparitinase (1 U/ml each) increased LDL binding by approximately 1.5-fold. Binding of lipoprotein (a) [Lp(a)] to both subendothelial matrix and Matrigel(R) increased 2-10-fold when the HSPG were removed by heparinase treatment. Incubation of endothelial cells with oxidized LDL (OxLDL) or lysolecithin resulted in decreased matrix proteoglycans and increased Lp(a) retention by matrix. The effect of OxLDL or lysolecithin on endothelial PG was abolished in the presence of HDL. The decrease in matrix HSPG was associated with production of a heparanase-like activity by OxLDL-stimulated endothelial cells. To test whether removal of HSPG exposes fibronectin, a candidate Lp(a) binding protein in the matrix, antifibronectin antibodies were used. The increased Lp(a) binding after HSPG removal was inhibited 60% by antifibronectin antibodies. Similarly, the increased Lp(a) binding to matrix from OxLDL-treated endothelial cells was inhibited by antifibronectin antibodies. We hypothesize that atherogenic lipoproteins stimulate endothelial cell production of heparanase. This enzyme reduces HSPG which in turn promotes Lp(a) retention.
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PMID:Subendothelial retention of lipoprotein (a). Evidence that reduced heparan sulfate promotes lipoprotein binding to subendothelial matrix. 925 86

Cell surface heparan sulfate proteoglycans facilitate uptake of growth-promoting polyamines (Belting, M., Persson, S., and Fransson, L.-A. (1999) Biochem. J. 338, 317-323; Belting, M., Borsig, L., Fuster, M. M., Brown, J. R., Persson, L., Fransson, L.-A., and Esko, J. D. (2001) Proc. Natl. Acad. Sci. U. S. A., in press). Here, we have analyzed the effect of polyamine deprivation on the structure and polyamine affinity of the heparan sulfate chains in various glypican-1 glycoforms synthesized by a transformed cell line (ECV 304). Heparan sulfate chains of glypican-1 were either cleaved with heparanase at sites embracing the highly modified regions or with nitrite at N-unsubstituted glucosamine residues. The products were separated and further degraded by heparin lyase to identify sulfated iduronic acid. Polyamine affinity was assessed by chromatography on agarose substituted with the polyamine spermine. In heparan sulfate made by cells with undisturbed endogenous polyamine synthesis, free amino groups were restricted to the unmodified, unsulfated segments, especially near the core protein. Spermine high affinity binding sites were located to the modified and highly sulfated segments that were released by heparanase. In cells with up-regulated polyamine uptake, heparan sulfate contained an increased number of clustered N-unsubstituted glucosamines and sulfated iduronic acid residues. This resulted in a greater number of NO/nitrite-sensitive cleavage sites near the potential spermine-binding sites. Endogenous degradation by heparanase and NO-derived nitrite in polyamine-deprived cells generated a separate pool of heparan sulfate oligosaccharides with an exceptionally high affinity for spermine. Spermine uptake in polyamine-deprived cells was reduced when NO/nitrite-generated degradation of heparan sulfate was inhibited. The results suggest a functional interplay between glypican recycling, NO/nitrite-generated heparan sulfate degradation, and polyamine uptake.
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PMID:Modulations of glypican-1 heparan sulfate structure by inhibition of endogenous polyamine synthesis. Mapping of spermine-binding sites and heparanase, heparin lyase, and nitric oxide/nitrite cleavage sites. 1157 85

During their migration into inflammatory sites, immune cells, such as T cells, secrete extracellular matrix (ECM)-degrading enzymes, such as heparanase, which, under mildly acidic conditions, degrade heparan sulfate proteoglycans (HSPG). We have previously shown that at pH 7.2, human placental heparanase loses its enzymatic activity, while retaining its ability to bind HSPG and promote T cell adhesion to unfractionated ECM. We now demonstrate that the 65-kDa recombinant human heparanase, which is devoid of enzymatic activity, but can still bind HSPG, captures T cells under shear flow conditions and mediates their rolling and arrest, in the absence or presence of stromal cell-derived factor 1 alpha (SDF-1 alpha; CXCL12), in an alpha(4)beta(1)-VCAM-1-dependent manner. Furthermore, heparanase binds to and induces T cell adhesion to key ECM components, like fibronectin and hyaluronic acid, in beta(1) integrin- and CD44-specific manners, respectively, via the activation of the protein kinase C and phosphatidylinositol 3-kinase intracellular signaling machineries. Although the nature of the putative T cell heparanase-binding moiety is unknown, it appears that heparanase exerts its proadhesive activity by interacting with the T cells' surface HSPG, because pretreatment of the cells with heparinase abolished their subsequent response to heparanase. Also, heparanase augmented the SDF-1 alpha-triggered phosphorylation of Pyk-2 and extracellular signal-regulated kinase-2 implicated in integrin functioning. Moreover, heparanase, which had no chemotactic effect on T cells on its own, augmented the SDF-1 alpha-induced T cell chemotaxis across fibronectin. These findings add another dimension to the known versatility of heparanase as a key regulator of T cell activities during inflammation, both in the context of the vasculature and at extravascular sites.
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PMID:Enzymatically quiescent heparanase augments T cell interactions with VCAM-1 and extracellular matrix components under versatile dynamic contexts. 1510 Feb 55

Histidine-rich glycoprotein (HRG) is an alpha2-glycoprotein found in mammalian plasma at high concentrations (approximately 150 microg/ml) and is distinguished by its high content of histidine and proline. Structurally, HRG is a modular protein consisting of an N-terminal cystatin-like domain (N1N2), a central histidine-rich region (HRR) flanked by proline-rich sequences, and a C-terminal domain. HRG binds to cell surfaces and numerous ligands such as plasminogen, fibrinogen, thrombospondin, C1q, heparin, and IgG, suggesting that it may act as an adaptor protein either by targeting ligands to cell surfaces or by cross-linking soluble ligands. Despite the suggested functional importance of HRG, the cell-binding characteristics of the molecule are poorly defined. In this study, HRG was shown to bind to most cell lines in a Zn(2+)-dependent manner, but failed to interact with the Chinese hamster ovary cell line pgsA-745, which lacks cell-surface glycosaminoglycans (GAGs). Subsequent treatment of GAG-positive Chinese hamster ovary cells with mammalian heparanase or bacterial heparinase III, but not chondroitinase ABC, abolished HRG binding. Furthermore, blocking studies with various GAG species indicated that only heparin was a potent inhibitor of HRG binding. These data suggest that heparan sulfate is the predominate cell-surface ligand for HRG and that mammalian heparanase is a potential regulator of HRG binding. Using recombinant forms of full-length HRG and the N-terminal N1N2 domain, it was shown that the N1N2 domain bound specifically to immobilized heparin and cell-surface heparan sulfate. In contrast, synthetic peptides corresponding to the Zn(2+)-binding HRR of HRG did not interact with cells. Furthermore, the binding of full-length HRG, but not the N1N2 domain, was greatly potentiated by physiological concentrations of Zn2+. Based on these data, we propose that the N1N2 domain binds to cell-surface heparan sulfate and that the interaction of Zn2+ with the HRR can indirectly enhance cell-surface binding.
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PMID:Histidine-rich glycoprotein binds to cell-surface heparan sulfate via its N-terminal domain following Zn2+ chelation. 1513 72

Heparanase is a mammalian endoglycosidase that degrades heparan sulfate (HS) at specific intrachain sites, an activity that is strongly implicated in cell dissemination associated with metastasis and inflammation. In addition to its structural role in extracellular matrix assembly and integrity, HS sequesters a multitude of polypeptides that reside in the extracellular matrix as a reservoir. A variety of growth factors, cytokines, chemokines, and enzymes can be released by heparanase activity and profoundly affect cell and tissue function. Thus, heparanase bioavailability, accessibility, and activity should be kept tightly regulated. We provide evidence that HS is not only a substrate for, but also a regulator of, heparanase. Addition of heparin or xylosides to cell cultures resulted in a pronounced accumulation of, heparanase in the culture medium, whereas sodium chlorate had no such effect. Moreover, cellular uptake of heparanase was markedly reduced in HS-deficient CHO-745 mutant cells, heparan sulfate proteoglycan-deficient HT-29 colon cancer cells, and heparinase-treated cells. We also studied the heparanase biosynthetic route and found that the half-life of the active enzyme is approximately 30 h. This and previous localization studies suggest that heparanase resides in the endosomal/lysosomal compartment for a relatively long period of time and is likely to play a role in the normal turnover of HS. Co-localization studies and cell fractionation following heparanase addition have identified syndecan family members as candidate molecules responsible for heparanase uptake, providing an efficient mechanism that limits extracellular accumulation and function of heparanase.
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PMID:Heparanase uptake is mediated by cell membrane heparan sulfate proteoglycans. 1529 2

The vascular endothelial growth factor (VEGF) family of proteins controls the formation and growth of blood vessels. The most potent and widely expressed isoform, VEGF165, is secreted as a disulfide-linked homodimer with two identical heparin-binding sites. Interactions with heparan sulfate (HS) regulate the diffusion, half-life, and affinity of VEGF165 for its signaling receptors. We have determined a number of key HS structural features that mediate the specific binding of the VEGF165 dimer. Carboxylate groups and 2-O-, 6-O-, and N-sulfation of HS contributed to the strength of the VEGF165 interaction; however, 6-O-sulfates appeared to be particularly important. Cleavage of HS by heparinase, heparitinase, or heparanase severely reduced VEGF165 binding. In contrast, K5 lyase-cleaved HS retained significant VEGF165 affinity, suggesting that binding sites for the growth factor are present within extended stretches of sulfation. Binding studies and molecular modeling demonstrated that an oligosaccharide 6 or 7 residues long was sufficient to fully occupy the heparin-binding site of a VEGF165 monomer. The data presented are consistent with a model whereby the two heparin-binding sites of the VEGF165 dimer interact simultaneously with highly sulfated S-domain regions of the HS chain that can be linked through a stretch of transition sequence.
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PMID:VEGF165-binding sites within heparan sulfate encompass two highly sulfated domains and can be liberated by K5 lyase. 1625 70

Heparanase has been previously associated with the metastatic potential, inflammation, and angiogenesis of tumor cells. Heparanase activity has been detected by means of UV absorption, radiolabeled substrates, electrophoretic migration, and heparan sulfate affinity assays. However, those methods have proven to be somewhat problematic with regards to application to actual biological samples, the accessibility of the immobilized substrates, experimental sensitivity, and the separation of degraded products. Rather than focusing on heparanase activity, then, we have developed a rapid, alternative colorimetric heparinase assay, on the basis of the recent finding that sulfated disaccharides generated from heparin by bacterial heparinase exhibit biological properties comparable to those from heparan sulfate by mammalian heparanase. In this study, the concentrations of porcine heparin and bacterial heparinase I were determined using a Sigma Diagnostics Kit. Morus alba was selected as a candidate through this assay system, and an inhibitor, resveratrol, was purified from its methanol extract. Its anti-metastatic effects on the pulmonary metastasis of murine B16 melanoma cells were also evaluated. Our findings suggest that this assay may prove useful as a diagnostic tool for heparinase inhibition, as an alternative anti-metastatic target.
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PMID:Colorimetric heparinase assay for alternative anti-metastatic activity. 1680 78


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