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)

Lipoprotein lipase (LPL), the rate limiting enzyme for hydrolysis of lipoprotein triglyceride, also mediates nonenzymatic interactions between lipoproteins and heparan sulfate proteoglycans. To determine whether cell surface LPL increases LDL binding to cells, bovine milk LPL was added to upregulated and nonupregulated human fibroblasts along with media containing LDL. LDL binding to cells was increased 2-10-fold, in a dose-dependent manner, by the addition of 0.5-10 micrograms/ml of LPL. The amount of LDL bound to the cells in the presence of LPL far exceeded the capacity for LDL binding via the LDL receptor. Treatment of fibroblasts with heparinase and heparitinase resulted in a 64% decrease in LPL-mediated LDL binding. Compared to studies performed without LPL, more LDL was internalized and degraded in the presence of LPL, but the time course was slower than that of classical lipoprotein receptor mediated pathways. In LDL receptor negative fibroblasts, LPL increased surface bound LDL > 140-fold, intracellular LDL > 40-fold, and LDL degradation > 6-fold. These effects were almost completely inhibited by heparin and anti-LPL monoclonal antibody. LPL also increased the binding and uptake by fibroblasts of apolipoprotein-free triglyceride emulsions; binding was increased > 8-fold and cellular uptake was increased > 40-fold with LPL. LPL increased LDL binding to THP-1 monocytes, and increased LDL uptake (4.5-fold) and LDL degradation (2.5-fold) by THP-1 macrophages. In the absence of added LPL, heparin and anti-LPL monoclonal antibodies decreased LDL degradation by > 40%, and triglyceride emulsion uptake by > 50%, suggesting that endogenously produced LPL mediated lipid particle uptake and degradation. We conclude that LPL increases lipid and lipoprotein uptake by cells via a pathway not involving the LDL receptor. This pathway may be important for lipid accumulation in LPL synthesizing cells.
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PMID:Lipoprotein lipase-mediated uptake and degradation of low density lipoproteins by fibroblasts and macrophages. 140 Oct 83

To study the interaction between low-density lipoprotein (LDL) and granules from rat serosal mast cells in vitro, mast cells were stimulated with the degranulating agent 48/80 to induce exocytosis of the secretory granules. Subsequent incubation of the exocytosed granules with 125I-LDL resulted in binding of the labelled LDL to the granules. When increasing amounts of agent 48/80 were added to mast-cell suspensions, a dose-dependent release of granules was observed and a parallel increase in the amount of 125I-LDL bound to granules resulted. 125I-LDL bound to a single class of high-affinity binding sites on the granules. At saturation, 105 ng of LDL were bound per microgram of granule protein. The lipoprotein binding to mast-cell granules was apolipoprotein(apo)-B + E-specific. Thus 125I-LDL binding to the granules was effectively compared for by LDL (apo-B) or by dimyristoyl phosphatidylcholine vesicles containing apo-E, but not by high-density lipoprotein (HDL3) containing apo-AI as their major protein component. Neutralization by acetylation of the positively charged amino groups of apo-B of LDL or presence of a high ionic strength in the incubation medium prevented LDL from binding to the granules, indicating the presence of ionic interactions between the positively charged amino acids of LDL and negatively charged groups of the granules. It could be demonstrated that LDL bound to the negatively charged heparin proteoglycan of the granules. Thus treatment of granules with heparinase resulted in loss of their ability to bind LDL, and substances known to bind to heparin, such as Toluidine Blue, avidin, lipoprotein lipase, fibronectin and protamine, all effectively competed with LDL for binding to the granules. The results show that LDL is efficiently bound to the heparin proteoglycan component of mast-cell granules once the mast cells are stimulated to release their granules into the extracellular space.
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PMID:Low-density-lipoprotein binding by mast-cell granules. Demonstration of binding of apolipoprotein B to heparin proteoglycan of exocytosed granules. 359 8

Bovine lactoferrin inhibits the clearance of remnant lipoproteins from the plasma and competes with the cell-surface binding of apolipoprotein (apo) E-enriched remnants. We established that lactoferrin inhibits remnant binding and uptake by interacting with both heparan sulfate proteoglycans (HSPG) and the low-density lipoprotein receptor-related protein (LRP). The binding of 125I-lactoferrin was inhibited 45% to 60% in HepG2 hepatocytes and wild-type Chinese hamster ovary (CHO) cells treated with heparinase to remove HSPG. In mutant CHO cells (pgsD-677) lacking HSPG, the level of 125I-lactoferrin binding was approximately 50% that seen with wild-type CHO cells; thus, about one half of lactoferrin binding appears to be mediated through cell-surface HSPG. A significant fraction of the residual binding of the lactoferrin appears to be mediated through the LRP. The 39-kd protein known to bind to the LRP and to block ligand interaction inhibited 125I-lactoferrin degradation in wild-type CHO cells by 60% to 65%. The addition of the 39-kd protein plus heparinase treatment reduced the binding by 85% to 90% (this combination blocks direct interaction with both the LRP and HSPG). However, it was also shown that the 39-kd protein bound to HSPG and the LRP. Heparinase treatment of wild-type CHO cells decreased the binding of the 125I-39-kd protein by approximately 40%, and the mutant CHO cells lacking HSPG bound half as much 125I-39-kd protein as wild-type CHO cells.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Lactoferrin binding to heparan sulfate proteoglycans and the LDL receptor-related protein. Further evidence supporting the importance of direct binding of remnant lipoproteins to HSPG. 752 99

Heparan sulfate proteoglycans (HSPG) are involved in the binding and uptake of apolipoprotein (apo) E-enriched remnant lipoproteins by cultured cells in vitro. To define the role of hepatic HSPG in remnant lipoprotein clearance in vivo, heparinase (30 units) was infused intravenously into mice to hydrolyze the liver HSPG and determine the effect of HSPG hydrolysis on remnant clearance by the liver. Liver HSPG were prelabeled by peritoneal injection of [35S]Na2SO4. Injection of heparinase decreased the amount of 35S-labeled liver HSPG by approximately 20-40% within 10-15 min. Heparinase infusion significantly inhibited the clearance of chylomicrons, chylomicron remnants, chylomicron remnants + apoE, rabbit beta-very low density lipoproteins (beta-VLDL), and beta-VLDL + apoE. Compared with saline injection in control mice, heparinase injection retarded the plasma clearance of the remnants by 1.5- to 2-fold and decreased liver uptake by 1.3- to 1.6-fold. Confocal fluorescence microscopy of thick slices of liver from mice injected with 1,1'-dioctadecyl-3,3,3', 3'-tetramethylindocarbocyanine-labeled beta-VLDL + apoE revealed markedly less intense fluorescence from hepatocytes in heparinase-treated animals compared with those in saline-treated control animals. Intravenous heparinase infusion did not inhibit the clearance of mouse low density lipoproteins (LDL), a ligand for the LDL receptor, and did not affect the clearance of alpha 2-macroglobulin, a ligand for the LDL receptor-related protein. The results suggest an important role of the liver HSPG in remnant clearance in vivo.
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PMID:Intravenous heparinase inhibits remnant lipoprotein clearance from the plasma and uptake by the liver: in vivo role of heparan sulfate proteoglycans. 753 27

Previously, we demonstrated in cultured dorsal root ganglion neurons that, in the presence of beta-migrating very low density lipoproteins (beta-VLDL), apolipoprotein (apo) E4, but not apoE3, suppresses neurite outgrowth. In the current studies, murine neuroblastoma cells (Neuro-2a) were stably transfected with human apoE3 or apoE4 cDNA, and the effect on neurite outgrowth was examined. The stably transfected cells secreted nanogram quantities of apoE (44-89 ng/mg of cell protein in 48 h). In the absence of lipoproteins, neurite outgrowth was similar in the apoE3- and apoE4-secreting cells. The apoE4-secreting cells, when incubated with beta-VLDL, VLDL, cerebrospinal fluid lipoproteins (d < 1.21 g/ml), or with triglyceride/phospholipid (2.7:1 (w/w)) emulsions, showed a reduction in the number of neurites/cell, a decrease in neurite branching, and an inhibition of neurite extension, whereas in the apoE3-secreting cells in the presence of a lipid source, neurite extension was increased. Uptake of beta-VLDL occurred to a similar extent in both the apoE3- and apoE4-secreting cells. With low density lipoproteins or with dimyristoylphosphatidylcholine emulsions, either alone or complexed with cholesterol, no differential effect on neurite outgrowth was observed. A slight differential effect was observed with apoE-containing high density lipoproteins. The differential effect of apoE3 and apoE4 in the presence of beta-VLDL was blocked by incubation of the cells with heparinase and chlorate, with lactoferrin, or with receptor-associated protein, all of which prevent the uptake of lipoproteins by the low density lipoprotein receptor-related protein (LRP). The data suggest that the secreted and/or cell surface-bound apoE interact with the lipoproteins and facilitate their internalization via the heparan sulfate proteoglycan-LRP pathway. The mechanism by which apoE3 and apoE4 exert differential effects on neurite outgrowth remains speculative. However, the data suggest that apoE4, which has been shown to be associated with late onset familial and sporadic Alzheimer's disease, may inhibit neuronal remodeling and contribute to the progression of the disease.
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PMID:Stable expression and secretion of apolipoproteins E3 and E4 in mouse neuroblastoma cells produces differential effects on neurite outgrowth. 759 57

An apolipoprotein (apo) E- and lipoprotein lipase-independent, high affinity, saturable and specific binding site and pathway for uptake of certain triglyceride-rich lipoproteins (TGRLP) by human monocyte-macrophages that leads to lipid accumulation and foam cell formation in vitro has been reported; two membrane binding activities were identified as receptor candidates with apparent molecular masses of 200 and 235 kDa [Gianturco et al. (1994) J. Lipid Res. 35, 1674-1687]. Here we present new evidence that these activities are TGRLP receptors with unique biochemical properties which distinguish them from other lipoprotein receptors. Protease and heparinase susceptibility studies demonstrate that (1) these activities have essential protein, but not heparan sulfate proteoglycan (HSPG) components; (2) the membrane binding proteins (MBPs) are located on the cell surface; (3) HSPGs do not facilitate TGRLP binding to this specific cellular site. Upon reduction, MBP 200 and 235 are both converted into a single, new binding activity of intermediate mobility (MBP 200R); all MBP forms displayed high affinity, saturable TGRLP binding with similar Kds (1.4-2.2 micrograms/mL). Notably, MBP 200R retained the combined ligand binding capacity of MBP 200 and 235 prior to reduction, demonstrating that, unlike members of the LDL receptor or the scavenger receptor families, disulfide bonds are not critical for activity. At 65 degrees C, MBP 235 was converted into MBP 200 without loss of total binding activity, suggesting heat dissociates a small subunit not required for binding from a common large protein subunit that binds TGRLP. Since the MBPs are found on the cell surface, are themselves functionally and structurally related, have distinctly different biochemical properties from members of the LDL receptor and scavenger receptor families, and share all critical characteristics with the cellular binding site, we hypothesize that they represent a new and unique receptor family for apoE- and lipoprotein lipase-independent uptake of TGRLP by human monocyte-macrophages.
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PMID:Human THP-1 monocyte-macrophage membrane binding proteins: distinct receptor(s) for triglyceride-rich lipoproteins. 761 11

Addition of apolipoprotein (apo) E to rabbit beta-very low density lipoproteins (beta-VLDL) has been shown to result in a marked enhancement of their binding and uptake by various cell types. Apolipoprotein E binds to lipoprotein receptors and proteoglycans. To distinguish between apoE binding to these sites, cells were treated with heparinase. Heparinase treatment of receptor-negative familial hypercholesterolemic (FH) fibroblasts and human hepatoma cells (HepG2) released 30-40% of newly synthesized cell surface 35S-labeled proteoglycans and decreased the binding of beta-VLDL+apoE to FH and normal fibroblasts and HepG2 cells by more than 80%. Furthermore, heparinase treatment significantly decreased the uptake of fluorescently labeled beta-VLDL+apoE by HepG2 cells and decreased cholesteryl ester synthesis in FH fibroblasts by 75%. Likewise, canine chylomicron remnants enriched in apoE demonstrated enhanced binding that was 80% inhibited by heparinase treatment of HepG2 cells. Heparinase treatment did not affect beta-VLDL (without added apoE) or low density lipoprotein (LDL) binding to these cells or the binding activity of beta-VLDL+apoE to the LDL receptor-related protein (LRP) or to the LDL receptor on ligand blots. Chinese hamster ovary (CHO) mutant cells lacking the synthesis of either heparan sulfate (pgsD-677) or all proteoglycans (pgsA-745) did not display any enhanced binding of the beta-VLDL+apoE. By comparison, wild-type CHO cells demonstrated enhanced binding of beta-VLDL+apoE that could be abolished by treatment with heparinase. These mutant cells and wild-type CHO cells possessed a similar amount of LRP, as determined by ligand blot analyses and by alpha 2-macroglobulin binding, and possessed a similar amount of LDL receptor activity, as determined by LDL binding. Therefore, we would interpret these data as showing that heparan sulfate proteoglycan may be involved in the initial binding of the apoE-enriched remnants with the subsequent involvement of the LRP in the uptake of these lipoproteins. It remains to be determined whether the heparan sulfate proteoglycan can function by itself in both the binding and internalization of the apoE-enriched remnants or whether the proteoglycan is part of a complex with LRP that mediates a two-step process, i.e. binding and subsequent internalization by the receptor.
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PMID:Role of heparan sulfate proteoglycans in the binding and uptake of apolipoprotein E-enriched remnant lipoproteins by cultured cells. 768 68

The low-density lipoprotein (LDL) receptor plays a crucial role in cholesterol metabolism. A related protein, designated the very low density lipoprotein (VLDL) receptor, that specifically binds apolipoprotein (apo) E has recently been characterized and shown to be expressed in heart, muscle and adipose tissue and the human monocyte-macrophage cell line THP-1. The VLDL receptor binds and internalizes VLDL and intermediate density lipoprotein from Watanabe heritable hyperlipidemic (WHHL) rabbits as well as beta-migrating VLDL from cholesterol-fed rabbits but not LDL from WHHL rabbits. Chinese hamster ovary (CHO) cells transfected with the rabbit VLDL receptor cDNA have now been shown to bind or internalize VLDL (d < 1.006 g/ml) isolated from fasted normolipidemic human subjects with lower affinity than WHHL-VLDL or rabbit beta-VLDL. However, binding and internalization were markedly enhanced when fasted human VLDL was preincubated with either recombinant human apoE (3/3) or lipoprotein lipase (LPL) in CHO cells overexpressing the rabbit or human VLDL receptor. CHO cells transfected with both the rabbit VLDL receptor cDNA and the human LPL cDNA effectively bound, internalized, and degraded fasted human VLDL without pretreatment. Treatment of heparinase reduced the effect of LPL-mediated binding at 4 degrees C, but the inhibitory effect was lower at 37 degrees C. Pseudomonas LPL also enhanced the binding of human fasted VLDL to the VLDL receptor at 37 degrees C in CHO cells overexpressing the human VLDL receptor. Taken together, LPL causes the enhancement of triglyceride-rich lipoproteins binding to the VLDL receptor via both the formation of bridge between lipoproteins and heparan sulfate proteoglycans and its lipolytic effect. Ligand blot analysis showed that the apparent molecular mass of the VLDL receptor is 118 kDa, which is smaller than that of the LDL receptor. These results indicate that the VLDL receptor recognizes both triglyceride-rich lipoproteins that are also relatively rich in apoE, as well as the remnants of triglyceride-rich lipoproteins after catabolism and the interaction with heparan sulfate proteoglycans by LPL. The VLDL receptor may thus function as a receptor for remnants of triglyceride-rich lipoproteins in extrahepatic tissues.
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PMID:Enhancement of the binding of triglyceride-rich lipoproteins to the very low density lipoprotein receptor by apolipoprotein E and lipoprotein lipase. 779 76

The initial step in the clearance of apolipoprotein (apo) E-enriched remnant lipoproteins from the plasma appears to be sequestration within the liver mediated by their binding to heparan sulfate proteoglycans (HSPG). The surface-bound remnants are believed to be internalized by their interaction with the low density lipoprotein (LDL) receptor-related protein or by the LDL receptor. Cholesterol-induced rabbit beta-very low density lipoproteins (beta-VLDL) enriched in human apoE3 display 4-5-fold enhanced binding to cultured cells. The present study attempts to determine whether recessive versus dominant type III hyperlipoproteinemia might be explained, at least in part, by a variable interaction of the mutant forms of apoE with the HSPG and impaired uptake. The beta-VLDL+apoE2(Arg158-->Cys), which is associated with recessive type III hyperlipoproteinemia, bound more poorly than beta-VLDL+apoE3 but still possessed significant enhanced binding (approximately 2-2.5-fold compared with beta-VLDL without added apoE) to HepG2 and McA-RH7777 cells. In comparison, beta-VLDL+apoE(Arg142-->Cys), beta-VLDL+apoE(Arg145-->Cys), and beta-VLDL+apoE-Leiden, which are associated with dominant type III hyperlipoproteinemia, bound more poorly. This same hierarchy of binding and uptake was determined by [14C]oleate incorporation into cholesteryl esters in LDL receptor-negative cells and by secretion of apoE3 and the variant apoE forms from McA-RH7777 cells. Furthermore, the enhanced binding of the apoE-enriched beta-VLDL was almost totally inhibited by heparinase treatment of the cells, and the basal binding activity was inhibited by 80-90% following addition of an LDL receptor antibody capable of blocking receptor-ligand interaction. The beta-VLDL enriched in apoE or apoE-dimyristoylphosphatidylcholine complexes bound to isolated HSPG from McA-RH7777 cells or the rat liver to a very similar degree. Likewise, the binding of beta-VLDL plus the various forms of apoE to the LDL receptor-related protein on ligand blots paralleled the results of other studies. In conclusion, all of the type III hyperlipoproteinemic apoE variants are defective in displaying enhanced binding to HSPG and in the cellular uptake initiated by HSPG. However, apoE2(Arg158-->Cys) displayed more activity than the variants associated with the dominant forms of type III hyperlipoproteinemia. The hierarchy of binding and uptake was as follows: apoE3 > apoE2(Arg158-->Cys) > apoE(Arg145-->Cys) > apoE(Arg142-->Cys) approximately apoE-Leiden (the latter two usually displaying very little, if any, enhanced binding and uptake). Thus, a correlation exists between the mode of expression of type III hyperlipoproteinemia and the binding and uptake of the specific apoE mutation.
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PMID:Variable heparan sulfate proteoglycan binding of apolipoprotein E variants may modulate the expression of type III hyperlipoproteinemia. 817 73

Rat hepatoma McA-RH7777 cells transfected with a human hepatic lipase (HL) cDNA synthesized and secreted 50-80 ng of human HL/mg of cell protein at 4 h, approximately 50% of which was bound to cell-surface heparan sulfate proteoglycans (HSPG). The newly synthesized HL possessed enzymatic activity. When rabbit beta-very low density lipoproteins (beta-VLDL) and canine chylomicrons or chylomicron remnants were incubated with HL-secreting cells, remnant binding and uptake were enhanced 3-fold compared with nontransfected cells. Furthermore, fluorescence microscopy showed enhanced uptake of 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine-labeled beta-VLDL by the HL-transfected cells. When 125I-beta-VLDL were added to conditioned medium from HL-secreting cells, the HL in the media enhanced the binding and uptake of the remnant lipoproteins by nontransfected cells about 3-fold. Likewise, surface-bound HL (without HL in the medium) also was able to mediate the enhanced binding of the remnants. This HL-enhanced binding was shown to be mediated by an interaction with cell-surface HSPG. Heparinase treatment to remove cell-surface HSPG or chlorate treatment to prevent HSPG sulfation of the HL-secreting cells abolished all the HL-mediated enhanced binding and uptake. Furthermore, heparinase pretreatment of nontransfected cells prevented the enhanced binding and uptake of beta-VLDL incubated with conditioned medium from HL-secreting cells. As binding was not enhanced in the absence of HSPG, an HL-HSPG initial interaction appears essential. Addition of apolipoprotein (apo) E to the beta-VLDL did not facilitate HL-mediated binding and uptake; in fact, beta-VLDL from apoE-null mice demonstrated a similar degree of enhanced binding as did rabbit beta-VLDL with or without added apoE. On the other hand, beta-VLDL from transgenic mice overexpressing binding-defective apoE(Arg142-->Cys) did not display any enhanced binding and uptake by the HL-secreting cells, and it appears that the apoE(Arg142-->Cys) actually inhibited the HL-mediated interaction. This mutant form of apoE is associated with a dominant mode of expression of type III hyperlipoproteinemia in contrast to the more commonly occurring recessive disorder. Impaired HL interaction with the apoE(Arg142-->Cys) beta-VLDL may contribute to remnant lipoprotein accumulation in the plasma of patients with this mutant form of apoE. Thus, HL contributes to the enhanced cell association of specific types of remnant lipoproteins by initiating their binding to cell-surface HSPG.
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PMID:Enhanced binding and uptake of remnant lipoproteins by hepatic lipase-secreting hepatoma cells in culture. 817 74


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