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)

Very low density lipoproteins ere isolated from plasma of swine by ultracentrifugal flotation. After delipidation, the lipid-free proteins were separated by chromatography on Sephadex G-150 AND DEAE-cellulose. A major apoprotein was isolated and shown to activate cows' milk lipoprotein lipase. Since human very low density lipoproteins also contain an activator protein, designated, apoC-II, we have called the pig protein, pig apoC-II. Pig apoC-II had a molecular weight of approximately 10 000 as determined by polyacrylamide gel electrophoresis in sodium dodecyl sulfate. The amino acid composistion showed the absence of histidine, cysteine and tryptophan; there was no evidence for carbohydrate. Treatment of pig apoC-II with carboxypeptidase indicated COOH-terminal serine. Rabbit antisera prepared to the pig protein gave single precipitin lines of complete identity to very low density lipoproteins, apoC-11. Using anti-pig apoC-II, a radioimmunoassay was developed which provides a convenient and reproducible method for measuring 5-1000 ng of apoprotein.
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PMID:Physical, chemical and immunochemical characterization of a lipoprotein lipase activator protein from pig plasma very low density lipoproteins. 18 30

The interaction of lipoprotein lipase (LpL) and its activator protein, apolipoprotein C-II (apoC-II), with a nonhydrolyzable phosphatidylcholine, 1,2-ditetradecyl-rac-glycero-3-phosphocholine (C14-ether-PC), was studied by fluorescence spectroscopy. A complex of 320 molecules of C14-ether-PC per LpL was isolated by density gradient ultracentrifugation in KBr. The intrinsic tryptophan fluorescence emission spectrum of LpL was shifted from 336 nm in the absence of lipid to 330 nm in the LpL-lipid complex; the shift was associated with a 40% increase in fluorescence intensity. Addition of C14-ether-PC vesicles to apoC-II caused a 2.5-fold increase in intrinsic tryptophan fluorescence and a shift in emission maximum from 340 to 317 nm. LpL and apoC-II/C14-ether-PC stoichiometries and binding constants were determined by measuring the increase in the intrinsic tryptophan fluorescence as a function of lipid and protein concentrations; for LpL the rate and magnitude of the fluorescence increases were relatively independent of temperature in the range 4-37 degrees C. A stoichiometry of 270 PC per LpL for the LpL-lipid complex compares favorably with the value obtained in the isolated complex. The dissociation constant (Kd) of the complex is 4.3 X 10(-8) M. For apoC-II, the stoichiometry of the complex is 18 PC per apoprotein, and the Kd is 3.0 X 10(-6) M. These data suggest that LpL binds more strongly than apoC-II to phosphatidylcholine interfaces.
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PMID:Interaction of lipoprotein lipase and apolipoprotein C-II with sonicated vesicles of 1,2-ditetradecylphosphatidylcholine: comparison of binding constants. 405 89

Synthetic fragments of apo-C-II, specifically labeled on their NH2-terminals with the 5-dimethylaminonaphthalene-1-sulfonyl (dansyl or DNS) fluorophore, have been prepared by solid phase peptide synthesis. When a complex is formed between bovine milk lipoprotein lipase and N-dansyl-apo-C-II peptides, resonance energy transfer occurs from the tryptophan residues of the enzyme to the dansyl-labeled peptides upon excitation at 280 nm. In the absence of lipid, the association constant increases 10-fold when the length of the DNS peptide is increased from apo-C-II-DNS(64-78) (0.04 X 10(6) M-1) to apo-C-II-DNS(60-78) (0.3 X 10(6) M-1). In the presence of lipid, the association constants are dependent on peptide chain length, and increase from 0.4 X 10(6) M-1 for apo-C-II-DNS(64-78) to 2.2 X 10(7) M-1 for apo-C-II-DNS(43-78). The interactions are specific for lipoprotein lipase, are disrupted by guanidinium chloride, are not affected by 1.0 M NaCl, and are competitive with the corresponding nondansylated peptide. Apolipoproteins C-III and A-I, at 5 to 1 molar ratios, had no effect on the interaction. These findings demonstrate the importance of the COOH-terminal region in the lipoprotein lipase-apo-C-II interaction and show that activation of the enzyme involves a specific protein-protein interaction.
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PMID:Interaction of synthetic N-5-dimethylaminonaphthalene-1-sulfonyl-apolipoprotein C-II peptides with lipoprotein lipase. 682 47

Lipoprotein lipase is the extrahepatic lipase responsible for the hydrolysis of triglycerides in chylomicrons and very low-density lipoproteins. Its enzymic activity and its location on the surface of endothelial cells are affected by the presence of fatty acids, implying that the protein possesses binding sites for that ligand. In this study, we examine the binding of fatty acids to LpL and describe factors that must be considered when the dissociation constant of the acceptor-ligand equilibrium is close to the critical micelle concentration of the fatty acid. The interaction of fatty acids with lipoprotein lipase (LpL) was studied by two methods. A new direct method, based on the LpL-induced increase in the apparent critical micelle concentration of the sodium soap of the fatty acid, indicates the presence of multiple high-affinity binding sites. In the second method, the specific binding of fatty acids to LpL was measured by quantitating the blue shift in the tryptophan fluorescence of LpL that occurs upon binding the ligand. Both methods suggest the existence of 4-6 fatty acid binding sites on LpL with a dissociation constant on the order of 10(-6)-10(-7) M. Further analysis of the blue shift indicates that at higher concentrations of fatty acid, large complexes are formed consisting of 260-310 molecules of fatty acid per LPL monomer. In contrast, no large complexes are formed with fatty acids that form crystals above their solubility limit.
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PMID:Interactions between fatty acids and lipoprotein lipase: specific binding and complex formation. 794 38

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

Very low (VLDL) and low density lipoproteins (LDL) were isolated from plasma of patients with the E3/3 phenotype which were divided into three groups based on their plasma triglyceride content: low (TG<200 mg/dl, TG(l)), intermediate (200<300 mg/dl, TG(i)300 mg/dl, TG(h)). The protein density (PD) on the VLDL and LDL surface was calculated from lipoprotein composition and protein location was studied by tryptophan fluorescence quenching by I(-) anions at 25 degrees C and 40 degrees C. A comparison of the TG(h) with the TG(l) group revealed a significant (<0.05) increase of the PD parameter as much as 21% for VLDL, but not for LDL where this parameter did not change for any group; generally, PD(LDL) values were 3.2-3.8-fold lower than PD(VLDL). In accordance with this difference, the tryptophan accessibility f in VLDL vs. LDL was lower at both temperatures. There were temperature-induced changes of the f parameter in opposite directions for these lipoproteins. The difference in f value gradually decreased for VLDL in the direction TG(l)TG(i)TG(h) while for LDL there was a U-shaped dependence for these groups. The Stern-Volmer quenching constant K(S-V) which is sensitive to both temperature and viscosity, did not change for VLDL, but K(S-V)(LDL) was 2-3-fold higher for the TG(i) group compared to the other two. The efficiencies of VLDL and LDL binding to the LDL receptor (LDLr) in vitro were compared by solid-phase assay free of steric hindrance observed in cell binding. The maximal number of binding sites did not change for either type of particles and between groups. The association constant K(a) and apolipoprotein (apo) E/apoB mole ratio values all increased significantly for VLDL, but not for LDL, in comparison of the TG(i+h) with the TG(l) group. Based on VLDL and LDL concentrations in serum and on the affinity constant values obtained in an in vitro assay, VLDL concentrations corresponding to 50% inhibition of LDL binding (IC(50)) were calculated in an assumption of the competition of both ligands for LDLr in vivo; the mean values of IC(50) decreased 2-fold when plasma TG exceeded 200 mg/dl. The functional dependences of K(a)(VLDL), IC(50) and apoE content in VLDL (both fractional and absolute) and in serum on TG content in the whole concentration range studied were fitted to a saturation model. For all five parameters, the mean half-maximum values TG(1/2) were in the range 52-103 mg/dl. The efficiency of protein-protein interactions is suggested to differ in normolipidemic vs. HTG-VLDL and apoE content and/or protein density on VLDL surface may be the primary determinant(s) of the increased binding of HTG-VLDL to the LDL receptor. ApoCs may compete with apoE for the binding to the VLDL lipid surface as plasma triglyceride content increases. The possible competition of VLDL with LDL for the catabolism site(s) in vivo, when plasma TG increases, could explain the atherogenic action of TG-rich lipoproteins. Moreover, the 'dual action' hypothesis on anti-atherogenic action of apoE-containing high density lipoproteins (HDL) in vivo is suggested: besides the well-known effect of HDL as cholesteryl ester catabolic outway, the formation of a transient complex of apoE-containing discs appearing at the site of VLDL TG hydrolysis by lipoprotein lipase with VLDL particles proposed in our preceding paper promotes the efficient uptake of TG-rich particles; in hypertriglyceridemia due to the diminished HDL content this uptake seems to be impaired which results in the increased accumulation of the remnants of TG-rich particles. This explains the observed increase in cholesterol and triglyceride content in VLDL and LDL, respectively, due to the CETP-mediated exchange of cholesteryl ester and triglyceride molecules between these particles.
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PMID:Structural peculiarities of the binding of very low density lipoproteins and low density lipoproteins to the LDL receptor in hypertriglyceridemia: role of apolipoprotein E. 1068 28

Apolipoprotein CII (apoCII) activates lipoprotein lipase (LPL). Seven residues, located on one face of a model alpha-helix spanning residues 59-75, are fully conserved in apoCII from ten different animal species. We have mutated these residues one by one. Substitution of Ala(59) by glycine, or Thr(62) and Gly(65) by alanine did not change the activation, indicating that these residues are outside the LPL-binding site. Replacement of Tyr(63), Ile(66), Asp(69), or Gln(70) by alanine lowered the affinity for LPL and the catalytic activity of the LPL-apoCII complex. For each residue several additional replacements were made. Most mutants retained some activating ability, but replacement of Tyr(63) by phenylalanine or tryptophan and Gln(70) by glutamate caused almost complete loss of activity. All mutants bound to liposomes with similar affinity as wild-type apoCII, and they also bound with similar affinity to LPL in the absence of hydrolyzable lipids. However, the inactive mutants did not compete with wild-type apoCII in the activation assay. Therefore, we conclude that the productive apoCII-LPL interaction may be dependent on substrate molecules. In summary, our data demonstrate that residues 63, 66, 69, and 70 are of special importance for the function of apoCII, but no single amino acid residue is absolutely crucial.
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PMID:Functional analyses of human apolipoprotein CII by site-directed mutagenesis: identification of residues important for activation of lipoprotein lipase. 1171 5

Eating and appetite disorders are frequent complications of the uremic syndrome which contribute to malnutrition in dialysis patients. The data suggest that uremic anorexia may occur with or without abdominal and visceral fat accumulation despite a lower food intake. This form of obesity (i.e., with low food intake and malnutrition) is more common in dialysis patients than obesity with high food intake. This article reviews the current knowledge regarding mechanisms responsible for appetite regulation in normal conditions and in uremic patients. Anorexia in dialysis patients has been historically considered as a sign of uremic toxicity due to "inadequate" dialysis as judged by uncertain means ("middle molecule" accumulation, Kt/V, "peak-concentration hypothesis," and others). We propose the tryptophan-serotonin hypothesis, based on a uremia-induced disorder in patients' amino acid profile--low concentrations of large neutral and branched-chain amino acids with high tryptophan levels. A high rate of tryptophan transport across the blood-brain barrier increases the synthesis of serotonin, a major appetite inhibitor. Inflammation may also play a role in the genesis of anorexia and malnutrition. For example, silent infection with Helicobacter pylori may be a source of cytokines with cachectic action; its eradication improves appetite and nutrition. The evaluation of appetite should take into account cultural and social aspects. Uremic patients showed a universal trend to carbohydrate preference and red meat refusal compared to healthy people. In contrast, white meat was less problematic. Uremic patients also have a remarkable attraction for citrics and strong flavors in general. Eating preferences or refusals have been related to the predominance of some appetite peptide modulators. High levels of cholecystokinin (CCK) (a powerful anorexigen) are associated with early satiety for carbohydrates and neuropeptide Y (NPY) (an orexigen) with repeated food intake. Obesity and elevated body mass index often falsely suggest a good nutritional status. In uremic patients (a hyperinsulinemia state), disorders in the regulation of fat distribution (insulin, leptin, insulin-like growth factor [IGF]-1, fatty acids, and disorders in receptors for insulin, lipoprotein lipase, mitochondrial uncoupling protein-2, and beta 3 adrenoreceptors) may cause abdominal fat accumulation without an increase in appetite. Finally, appetite regulation in uremia is highly complex. Disorders in adipose tissue, gastrointestinal and neuropeptides, retained or hyperproduced inflammatory end products, and central nervous system changes may all play a role. Uremic anorexia may be explained by a hypothalamic hyperserotoninergic state derived from a high concentration of tryptophan and low branched-chain amino acids.
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PMID:Eating behavior disorders in uremia: a question of balance in appetite regulation. 1471 11

Tumor growth and abnormal cell survival were shown to be associated with a number of cellular metabolic abnormalities revealed by impaired oral glucose tolerance, depressed lipoprotein lipase activity leading to hypertriglyceridemia, and changes in amino acid profile as evidenced by increased plasma free tryptophan levels in patients with breast, lung, colon, stomach, and other cancers from various origins. The above findings seem to relate to or indicate a shift to non-oxidative metabolic pathways in cancer. In contrast to normal cells, cancer cells may lose the ability to utilize aerobic respiration due to either defective mitochondria or hypoxia within the tumor microenvironments. Glucose was shown to be the major energy source in cancer cells where it utilizes aerobic /anaerobic glycolysis with the resultant lactic acid formation. The role of energetic modulations and use of glycolytic inhibitors on cancer / normal cell survival is not clearly established in the literature. Therefore, the purpose of this study was to evaluate six glycolytic inhibitors namely, sodium ascorbate, oxalic acid, oxaloacetic acid, sodium citrate, fructose diphosphate (FDP) and sodium bicarbonate at microM concentrations on growing A549 (lung cancer) and MRC-5 (normal; human lung fibroblast) cell lines with the objective of determining their influence on cell survival. Exposed and non-exposed cells were tested with phase contrast micro scanning, survival / death and metabolic activity trends through MTT-assays, as well as death end-point determinations by testing re-growth on complete media. Results showed that oxalic acid and oxaloacetic acid both influenced the pH of the medium and resulted in differential massive cell debris within the exposure period. Sodium ascorbate, sodium citrate, sodium bicarbonate and FDP did not cause pH changes; however, they caused detectable cell disfigurement and loss of metabolic activity and survival/ death end points with the resultant death of the A549 cell line. MRC-5 cells were differentially unaffected by exposure to sodium ascorbate, sodium citrate, sodium bicarbonate, and oxaloacxetic acid, underwent complete recovery and remained both attached and healthy for 6 weeks upon subculture when transferred to a new complete medium. Oxalic acid did not show differential modulation with the consequent loss of survival and death of the MRC-5 cell line. These studies show the potential for exploiting cellular metabolic differences in cancer control.
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PMID:Differential modulation of intracellular energetics in A549 and MRC-5 cells. 1748 66

Non-alcoholic fatty liver disease (NAFLD) is one of the most prevalent chronic liver diseases worldwide with an unclear mechanism. Long non-coding RNAs (lncRNAs) have recently emerged as important regulatory molecules. To better understand NAFLD pathogenesis, lncRNA and messenger RNA (mRNA) microarrays were conducted in an NAFLD rodent model. Potential target genes of significantly changed lncRNA were predicted using cis/trans-regulatory algorithms. Gene Ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were then performed to explore their function. In the current analysis, 89 upregulated and 177 downregulated mRNAs were identified, together with 291 deregulated lncRNAs. Bioinformatic analysis of these RNAs has categorized these RNAs into pathways including arachidonic acid metabolism, circadian rhythm, linoleic acid metabolism, peroxisome proliferator-activated receptor (PPAR) signaling pathway, sphingolipid metabolism, steroid biosynthesis, tryptophan metabolism and tyrosine metabolism were compromised. Quantitative polymerase chain reaction (qPCR) of representative nine mRNAs and eight lncRNAs (named fatty liver-related lncRNA, FLRL) was conducted and this verified previous microarray results. Several lncRNAs, such as FLRL1, FLRL6 and FLRL2 demonstrated to be involved in circadian rhythm targeting period circadian clock 3 (Per3), Per2 and aryl hydrocarbon receptor nuclear translocator-like (Arntl), respectively. While FLRL8, FLRL3 and FLRL7 showed a potential role in PPAR signaling pathway through interaction with fatty acid binding protein 5 (Fabp5), lipoprotein lipase (Lpl) and fatty acid desaturase 2 (Fads2). Functional experiments showed that interfering of lncRNA FLRL2 expression affected the expression of predicted target, circadian rhythm gene Arntl. Moreover, both FLRL2 and Arntl were downregulated in the NAFLD cellular model. The current study identified lncRNA and corresponding mRNA in NAFLD, providing new insight into the pathogenesis of NAFLD. Moreover, we identified a new lncRNA FLRL2, that might participate NAFLD pathogenesis mediated by Arntl.
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PMID:Long Non-Coding RNA Profiling in a Non-Alcoholic Fatty Liver Disease Rodent Model: New Insight into Pathogenesis. 2827 12


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