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:3.1.1.34 (lipoprotein lipase)
7,025 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Synthesis and secretion of lipoprotein lipase was studied in two mutants of Chinese hamster ovary (CHO) cells which, due to a lack of xylosyl transferase (pgsA-745) or galactosyl transferase (pgsB-761), respectively, were deficient in heparan sulfate and chondroitin sulfate. One of the mutants (pgsB-761) was two- to threefold more active in synthesis and secretion of catalytically active lipoprotein lipase than the other mutant, which was about as active as the wild-type (K1) cells. A similar relation was found when lipoprotein lipase was metabolically labelled with 35S-methionine and then immunoprecipitated. Heparin stimulated secretion from all three cell types to a similar extent (about twofold). Heparin-releasable binding of 125I-labelled lipoprotein lipase was lower to either of the mutant cells than to the wild-type cells. Binding to the wild-type cells was reduced by heparitinase, while the low binding to the mutants was not affected. By immunogold labelling of cryosections, lipoprotein lipase was detected on the plasma membranes and on the inside of secretory vesicles of both wild-type and mutant cells, suggesting that some carrier could be involved. Inhibition of vesicular transport by monensin caused accumulation of lipoprotein lipase in the cells. In wild-type cells the lipase was mainly on the inside of vesicular structures, while in the mutants the main part was associated with membranous bodies that formed within the vesicles during a chase period. These results suggest that if lipoprotein lipase needs a carrier during intracellular assembly and transport, this function can be fulfilled by some structure other than heparan sulfate.
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PMID:Synthesis and secretion of lipoprotein lipase in heparan sulfate-deficient Chinese hamster ovary cells. 868 48

We examined the structure-function relationship of human lipoprotein lipase (hLPL) in its ability to enhance the binding and catabolism of very low density lipoproteins (VLDL) in COS cells. Untransfected COS cells did not bind to or catabolize normal VLDL. Expression of wild-type hLPL by transient transfection enhanced binding, uptake, and degradation of the VLDL (a property of LPL that we call bridge function). Heparin pretreatment and a monoclonal antibody ID7 that blocks LDL receptor-binding domain of apoE both inhibited binding, and apoE2/E2 VLDL from a Type III hyperlipidemic subject did not bind. However, LDL did not reduce 125I-VLDL binding to the hLPL-expressing cells, whereas rabbit beta-VLDL was an effective competitor. By contrast, LDL reduced uptake and degradation of 125I-VLDL to the same extent as excess unlabeled VLDL or beta-VLDL. These data suggest that binding occurs by direct interaction of VLDL with LPL but the subsequent catabolism of the VLDL is mediated by the LDL receptor. Mutant hLPLs that were catalytically inactive, S132A, S132D, as well as the partially active mutant, S251T, and S172G, gave normal enhancement of VLDL binding and catabolism, whereas the partially active mutant S172D had markedly impaired capacity for the process; thus, there is no correlation between bridge function and lipolytic activity. A naturally occurring genetic variant hLPL, S447-->Ter, has normal bridge function. The catalytic center of LPL is covered by a 21-amino acid loop that must be repositioned before a lipid substrate can gain access to the active site for catalysis. We studied three hLPL loop mutants (LPL-cH, an enzymatically active mutant with the loop replaced by a hepatic lipase loop; LPL-cP, an enzymatically inactive mutant with the loop replaced by a pancreatic lipase loop; and C216S/C239S, an enzymatically inactive mutant with the pair of Cys residues delimiting the loop substituted by Ser residues) and a control double Cys mutant, C418S/C438S. Two of the loop mutants (LPL-cH and LPL-cP) and the control double Cys mutant C418S/C438S gave normal enhancement of VLDL binding and catabolism, whereas the third loop mutant, C216S/C239S, was completely inactive. We conclude that although catalytic activity and the actual primary sequence of the loop of LPL are relatively unimportant (wild-type LPL loop and pancreatic lipase loops have little sequence similarity), the intact folding of the loop, flanked by disulfide bonds, must be maintained for LPL to express its bridge function.
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PMID:Structure-function relationship of lipoprotein lipase-mediated enhancement of very low density lipoprotein binding and catabolism by the low density lipoprotein receptor. Functional importance of a properly folded surface loop covering the catalytic center. 870 93

Heparin can cause an artifactual elevation in the concentration of unbound (free) thyroxine (T4) in the plasma, particularly when measured by equilibrium dialysis. The lipase released into the plasma by heparin acts on substrate (triglycerides; TG) in the plasma in vitro to release nonesterified (free) fatty acids (FFA), which, in high concentrations, inhibit the binding of T4 to its plasma binding proteins. This artifact occurs only in the presence of sufficient substrate (serum TG greater than approximately 180 mg/dL), and is most pronounced in methods requiring long incubation times. We observed this artifact in a patient receiving intralipid and subcutaneous (sc) heparin. Plasma-free T4, when measured by equilibrium dialysis, was elevated, but was normalized when the in vitro generation of FFA during equilibrium dialysis was prevented by prior treatment of the sample with protamine to inhibit lipoprotein lipase and with an antibody to hepatic triglyceride lipase. This observation caused us to investigate formally whether heparin, at standard sc doses or at iv doses even lower than those that are commonly used to flush iv lines (100-300 U), could also cause this artifact. We gave increasing doses of heparin at weekly intervals to each of three normal volunteers and measured FFA generation in their plasma (supplemented with 250 mg/dL triglycerides) under conditions simulating equilibrium dialysis. We found that, indeed, iv doses of heparin as low as 0.08 U/kg (5.6 U in a 70-kg subject) as well as a standard dose of sc heparin (5000 U) could release significant lipase activity into the plasma and, in the setting of sufficient substrate, cause enough in vitro generation of FFA to artifactually increase the serum-free T4 concentration when measured by equilibrium dialysis. These results indicate that equilibrium dialysis may not always be the best method for assessing serum-free T4 concentrations in hospitalized patients, and should be taken into account when interpreting previous studies demonstrating inhibitors of T4-serum protein binding in sera from hospitalized patients.
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PMID:Extremely low doses of heparin release lipase activity into the plasma and can thereby cause artifactual elevations in the serum-free thyroxine concentration as measured by equilibrium dialysis. 873 76

Glycosylation of lipoprotein lipase (LPL) was studied in human subcutaneous lipomas. Heparin-releasable LPL activities were higher in lipomas than those in adjacent normal adipose tissues, and showed good correlation with cellular LPL protein mass. Molecular weight of LPL subunit was 57 kDa in both tissues. After endoglycosidase H-digestion, two types of LPL subunits were found in normal adipose tissues; partially sensitive (55 kDa) and totally sensitive (52 kDa) form. In lipoma tissues, the fraction of partially sensitive form (55 kDa) was increased comparing with control adipose tissues. These results suggest that partially sensitive subunits constitute the major secretable form of LPL in human subcutaneous lipomas.
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PMID:Glycosylation of lipoprotein lipase in human subcutaneous lipomas. 882 Sep 86

Chronic renal failure (CRF) is associated with hypertriglyceridemia, impaired clearance of very low density lipoproteins (VLDL) and chylomicrons and their remnants as well as triglyceride-enrichment of various lipoproteins. These abnormalities are indicative of depressed lipoprotein lipase (LPL)-mediated hydrolysis of triglycerides in VLD and chylomicrons. In fact, impaired post-heparin lipolytic activity and decreased adipose tissue LPL activity has been previously demonstrated in CRF. The reduction in LPL activity in CRF has been attributed to PTH-induced insulin resistance and the presence of excess lipase inhibitors in uremic plasma. However, the effect of CRF on gene expression of LPL has not been elucidated and was studied here. Heparin-releasable, detergent-extractable and total LPL activities, as well as LPL mRNA of the heart, soleus muscle and fat body were determined in male Sprague-Dawley rats at baseline and on weeks 1, 3 and 6 following 5/6 nephrectomy (CRF group) or sham operation (control group). The CRF group exhibited a marked and steady rise in plasma triglycerides along with a steady decline in LPL activities and mRNA levels of all tissues studied. In contrast, the study parameters remained virtually unchanged throughout the study period in the control group. A strong inverse correlation was found between plasma triglycerides and LPL activity in the study animals. LPL activity was directly related to LPL mRNA. We conclude that CRF results in marked down-regulation of LPL expression that can contribute to dyslipidemia and altered energy metabolism in uremia. The effect of depressed LPL expression is compounded by the previously demonstrated elevations in uremic plasma of Apo C-III and pre-beta-HDL, which are potent inhibitors of LPL.
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PMID:Down-regulation of tissue lipoprotein lipase expression in experimental chronic renal failure. 894 76

Standard heparin as well as low molecular weight heparin (LMWH) increase lipid levels in serum. It has been reported that a diet rich in long chain saturated fatty acids can enhance the susceptibility to experimental thrombosis. The mechanism by which serum fatty acids may provoke thrombosis is not clear. It is possible that the fatty acids change the properties of the cell membrane and thereby modify the response of platelets to aggregating agents. Heparin and its LMW fractions, by mobilising lipoprotein lipase that hydrolyses serum triglycerides (TG), cause the serum TG to increase, a well known "clearing effect' of heparin in turbid lipemic plasma. This effect may have no significance when it lasts for a short time; however, a long-lasting heparin effect on TG serum levels may have important consequences. The purpose of this study was to examine the time span of the action of heparin and its fractions and to investigate variations in the concentration of digoxin, which is a compound with narrow therapeutic width. The investigated substances after 2 days administration, provoked serum concentration increases of free fatty acids (FFA), TG and HDL-C. Seven days after stopping drug administration, FFA and HDL-C levels remained high, while triglycerides declined. Serum total cholesterol remained unchanged throughout. Digoxin levels increased non-significantly after heparin administration and during swimming stress, while a lipid diet caused a serum digoxin concentration increase.
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PMID:How harmless is FFA enhancement? 898 Sep 17

Previous studies had pointed to an important function of a putative exposed loop in the C-terminal domain of lipoprotein lipase for activity against emulsified lipid substrates. This loop contains 3 tryptophan residues (Trp390, Trp393, and Trp394). We have expressed and characterized lipase mutants with tryptophan to alanine substitutions at positions 55, 114, 382, 390, 393, and 394 and a double mutant at residues 393 and 394. The substitutions in the N-terminal domain (W55A and W114A) led to poor expression of completely inactive lipase variants. Heparin-Sepharose chromatography showed that mutant W114A eluted at the same salt concentration as inactive wild-type monomers, indicating that this substitution prevented subunit interaction or led to an unstable dimer. In contrast, all mutants in the C-terminal domain were expressed as mixtures of monomers and dimers similarly to the wild-type. The dimers displayed at least some catalytic activity and had the same apparent heparin affinity as the active wild-type dimers. The mutants W390A, W393A, W394A, and W393A/W394A had decreased reactivity with the monoclonal antibody 5D2, indicating that the 5D2 epitope is longer than was reported earlier, or that conformational changes affecting the epitope had occurred. The mutants W390A, W393A, W394A, and W393A/W394A had decreased catalytic activity against a synthetic lipid emulsion of long-chain triacylglycerols (IntralipidR) and in particular against rat lymph chylomicrons. The most pronounced decrease of activity was found for the double mutant W393A/W394A which retained only 6% of the activity of the wild-type lipase, while 70% of the activity against water-soluble tributyrylglycerol was retained. In the case of chylomicrons also the affinity for the substrate particles was lowered, as indicated by severalfold higher apparent Km values. This effect was less prominent with the synthetic lipid emulsion. We conclude that the tryptophan cluster Trp390-Trp393-Trp394 contributes to binding of lipoprotein lipase to lipid/water interfaces. Utilizing different lipid substrates in different physical states, we have demonstrated that the tryptophan residues in the C-terminal domain may have a role also in the productive orientation of the enzyme at the lipid/water interface.
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PMID:Mutation of tryptophan residues in lipoprotein lipase. Effects on stability, immunoreactivity, and catalytic properties. 899 62

Interaction of different classes of lipoproteins with heparan sulfate, heparin, and lipoprotein lipase was studied by a surface plasmon resonance based technique on a BIAcore. The proteoglycans were covalently attached to sensor chips as previously described [Lookene, A., Chevreuil, O., Ostergaard, P., & Olivecrona, G. (1996) Biochemistry 35, 12155-12163]. Binding of all lipoproteins, except for beta-VLDL, to endothelial heparan sulfate was low. Binding of chylomicrons (from rat lymph) and of human VLDL was much increased by the presence of lipoprotein lipase. With human LDL, binding was low in the absence of lipase or at low lipase concentrations. For efficient binding, 2-4 lipase dimers per LDL particle were necessary, indicating cooperativity in the interaction. In contrast, HDL did not bind under any conditions. Heparin had higher binding capacity for lipoproteins than heparan sulfate. This was due to a higher number of binding sites on the heparin chains. Binding of LDL, VLDL, and chylomicrons to heparan sulfate-covered surfaces, both in the presence and in the absence of lipoprotein lipase, was characterized by high values for association rate constants (10(4)-10(5) M(-1) s(-1)) and low values for dissociation rate constants (10(-4)-10(-5) M(-1) s(-1)). In some experiments, rabbit beta-VLDL were directly immobilized to the sensor chips. Binding of lipoprotein lipase to these surfaces was characterized by a very high association rate constant (10(6) M(-1) s(-1)). The dissociation of triacylglycerol-rich lipoproteins was more rapid with catalytically active lipase than with active site-inhibited lipase. It was also markedly increased in the presence of free heparin, suggesting fast exchange kinetics at the surface. Based on that, we propose that lipoproteins are relatively mobile at heparan sulfate covered surfaces. Our study emphasizes the important role of lipoprotein lipase, or molecules with similar properties (apolipoprotein E, hepatic lipase), as mediators for binding of lipoproteins to proteoglycans. It also demonstrates the great potential for the use of biosensors for studies of lipoprotein interactions.
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PMID:Interaction of lipoproteins with heparan sulfate proteoglycans and with lipoprotein lipase. Studies by surface plasmon resonance technique. 913 89

Heparin administration to diabetic rats caused no change in VLDL, an increase in IDL and a decrease in LDL on electrophoretic analysis of plasma lipoproteins, while the administration to control rats markedly decreased VLDL and increased IDL and LDL. Both hepatic triglyceride lipase (HTGL) and lipoprotein lipase (LPL) activities in the postheparin plasma were lower in the diabetic rats than in the controls, and the reduction of HTGL activity was greater than that of LPL activity in the diabetic rats. The LPL activity in the adipose tissue was lower in the diabetic rats than in the controls, but the activities in the cardiac and skeletal muscles were similar in the two rats. The HTGL-catalyzed fatty acid (FA) releases from the diabetic VLDL and IDL were lower than those from the normal rat VLDL and IDL, while the LPL-catalyzed FA release in the diabetic rats was not different from those in the controls. The decreases in LPL and HTGL activities and the markedly impaired susceptibility of IDL to HTGL coincide well with the postheparin changes in plasma lipoproteins in diabetic rats, an increase in IDL and a decrease in LDL.
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PMID:Lipase activities in post-heparin plasma and tissues, and susceptibilities of lipoproteins in experimental diabetic rats. 922 15

Vascular endothelial-bound lipoprotein lipase (LPL), also known as heparin-releasable LPL, catalyzes the breakdown of the triglyceride component of lipoproteins and is rate-limiting for free fatty acid transport to tissues. We previously demonstrated that heparin-releasable LPL activity increases in diabetic Wistar rat hearts, whereas with the development of hypertension in spontaneously hypertensive rats (SHR), there is a concomitant and progressive reduction in LPL activity. The objective of the present study was to examine the regulation of cardiac LPL activity in SHR-diabetic rats. Heparin perfusion of the isolated Langendorff heart induced the release of LPL activity. SHR hearts demonstrated a reduction in peak heparin-releasable LPL activity, relative to Wistar controls. However, induction of streptozotocin-induced diabetes in SHR, as in Wistar rats, increased peak heparin-releasable LPL activity in perfused hearts. The elevated heparin-releasable LPL peak could not be accounted for by enhanced LPL synthesis in that both cellular and surface-bound LPL activities in myocytes from SHR-diabetic rats were low relative to control. Chronic (12-day) insulin treatment of SHR-diabetic rats reduced the augmented heparin-releasable LPL activity and increased cell-associated LPL activity. Moreover, acute (90-minute) treatment of SHR-diabetic rats with rapid-acting insulin also reduced the heparin-releasable LPL activity to normal, although it had no effect on the low cellular LPL activity. These results demonstrate that the diabetes-induced augmentation of cardiac LPL counteracts the reduction in enzyme activity associated with hypertension. This may serve to increase the delivery of free fatty acid to the heart, and the resultant metabolic changes may lead to the severe cardiomyopathy observed in the hypertensive-diabetic rat heart.
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PMID:Streptozotocin-induced diabetes enhances cardiac heparin-releasable lipoprotein lipase activity in spontaneously hypertensive rats. 949 76


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