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)

Apolipoprotein C-II (apoC-II), a protein constituent of very low density lipoproteins of human plasma and the activator protein of lipoprotein lipase, has been isolated and its amino acid sequence has been studied. The protein has 78 amino acid residues and is lacking cysteine, cystine, and histidine. Chromatography on Bio-Gel P-30 in 25% formic acid of the cyanogen bromide digest of apoC-II yields three fragments designated as CNBr-I, -II, and -III. They contained 50, 19, and 9 residues, respectively. The alignment of the cyanogen bromide fragments has been established as CNBr-III-I-II by isolation and sequence of the tryptic peptides of the intact protein. The amino acid sequences of the tryptic and CNBr peptides were determined by conventional methods. With this information, it was possible to establish the complete amino acid sequence of apoC-II.
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PMID:Primary structure of very low density apolipoprotein C-II of human plasma. 19 44

Apolipoprotein C-II (apoC-II), a protein constituent of human very low density lipoproteins, is the activator for lipoprotein lipase (LPL; triacylglycerol acyl-hydrolase, EC 3.1.1.3). The amino acid sequence of the 78 residues of apoC-II has recently been established in this laboratory. To determine the minimal sequence requirements for activation, we have prepared both native and synthetic fragments of apoC-II and tested them for their ability to activate LPL. Cyanogen bromide fragments of apoC-II corresponding to residues 1--9 and 10--59 had little ability to activate LPL. However, the COOH-terminal cyanogen bromide fragment corresponding to residues 60--78 increased hydrolysis 4-fold compared to an average of 9-fold activation for the same concentration of apoC-II. The synthetic peptide containing residues 60--78 prepared by solid-phase techniques enhanced the lipolysis 3-fold. Addition of five residues produced a synthetic fragment 55--78 that enhanced the release of fatty acid 12-fold compared to 13-fold for intact apoC-II. By contrast, the synthetic peptide containing residues 66--78 did not activate. Removal of the three COOH-terminal residues, Gly-Glu-Glu, from fragment 60--78 decreased the ability to activate LPL by greater than 95%. These studies suggest that the maximal activation of LPL by apoC-II requires a minimal sequence contained within residues 55--78.
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PMID:Activation of lipoprotein lipase by native and synthetic fragments of human plasma apolipoprotein C-II. 27 Jul 15

The substrate specificity of a calcium-independent, 97-kDa phospholipase B purified from guinea pig intestine was further investigated using various natural and synthetic lipids. The enzyme was equally active toward enantiomeric phosphatidylcholines under conditions allowing a strict phospholipase A activity. The lysophospholipase activity declined with the following substrates: 1-acyl-sn-glycero-3-phosphocholine greater than 1-palmitoyl-propanediol-3-phosphocholine greater than 1-palmitoyl-glycol-2-phosphocholine, suggesting some influence of the polar residue vicinal to the cleavage site. The enzyme also acted on various neutral lipids including triacylglycerol, diacylglycerol, and monoacylglycerol, whereas cholesteryl oleate remained refractory to enzymatic hydrolysis. The lipase hydrolyzed sequentially the sn-2 and sn-1 acyl ester bonds of diacylglycerol, although some direct cleavage of the external acyl ester bond could also occur, as shown with diacylglycerol analogues bearing a nonhydrolyzable alkyl ether or amide bond in the sn-1 or sn-2 position. The three main activities of the enzyme (phospholipase A2, lysophospholipase, and diacylglycerol lipase) were resistant to 4-bromophenacyl bromide, but they were inhibited by N-ethylmaleimide, 5,5'-dithiobis-(2-nitrobenzoic acid), and diisopropyl fluorophosphate, suggesting the possible involvement of both cysteine and serine residues in a single active site. It is concluded that guinea pig intestinal phospholipase B, which was also detected in rat and rabbit, is actually a glycerol ester lipase with broad substrate specificity and some unique enzymatic properties.
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PMID:Substrate specificity of phospholipase B from guinea pig intestine. A glycerol ester lipase with broad specificity. 161 44

The primary structure of bovine milk lipoprotein lipase (bLPL) was determined by alignment of peptides produced by tryptic digestion, Staphylococcus aureus V8 protease digestion, and cyanogen bromide cleavage. bLPL consists of 450 amino acid residues. Most tryptic peptides were isolated and analyzed, except for the dipeptide, Glu-Lys (position 423-424), and the 2 Lys at positions 416 and 488. Peptides resulting from digestion by S. aureus V8 protease and cyanogen bromide cleavage filled the missing part and completed the primary sequence of bLPL. The NH2 terminus of bLPL was determined to be Asp by sequencing the intact protein with a gas phase sequencer for up to 30 residues, whereas the COOH terminus was identified as Gly through, carboxyl peptidase Y cleavage. The enzyme contains 10 cysteine residues, all of which exist in disulfide linkages. They are formed between Cys29 and Cys42, Cys218 and Cys241, Cys266 and Cys285, Cys277, and Cys280, and Cys420 and Cys440. The sites of N-glycosylation were identified at Asn44 and Asn361. In accordance with a common structural homology of serine-type esterases, -G-X-S-X-G- (Yang, C. Y., Manoogian, D., Pao, Q., Lee, F., Knapp, R. D., Gotto, A. M., Jr., and Pownall, H. J. (1987) J. Biol. Chem., 262, 3086-3191), the active site serine of bLPL was assigned to the serine at position 134. The chymotrypsin nick of bLPL was determined to be between residues 390 and 391. A model of the enzyme is proposed on the basis of our data and available chemical data.
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PMID:Structure of bovine milk lipoprotein lipase. 267 42

The recently identified diacylglycerol lipase activity in membranes of chromaffin cells from bovine adrenal medulla [24] is now shown to consist of two enzymes working in series. First the predominantly saturated fatty acid in the sn-1-position is split by a diacylglycerol lipase (glycerol ester hydrolase, EC 3.1.1.34). Subsequently the resulting sn-2-monoacylglycerol is split by a monoacylglycerol lipase (glycerol-monoester acylhydrolase, EC 3.1.1.23) which prefers sn-2-arachidonoyl-monoacylglycerol to sn-2-palmitoyl-monoacylglycerol. At pH 4.0 only the diacylglycerol lipase is active, whereas the monoacylglycerol lipase is irreversibly inactivated. At pH 6.0 both enzymes are active. Pretreatment of the membranes at pH 10 leads to the selective inactivation of the diacylglycerol lipase. Both enzymes are Ca2+- and calmodulin-independent and both are partially inhibited by p-bromophenacyl bromide, however, only at relatively high concentrations of the inhibitor. Chlorpromazine inhibits the diacylglycerol lipase to about the same extent as p-bromophenacyl bromide but the monoacylglycerol lipase is less sensitive. The specific diacylglycerol lipase inhibitor RHC 80267 (1,6-di(O-(carbamoyl)cyclohexanone oxime)hexane) only interacts with the first step, i.e. the diacylglycerol lipase.
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PMID:Diacylglycerol breakdown in plasma membranes of bovine chromaffin cells is a two-step mechanism mediated by a diacylglycerol lipase and a monoacylglycerol lipase. 368 85

Human lipoprotein lipase and hepatic triglyceride lipase were purified to homogeneity from post-heparin plasma. These enzymes were purified 250,000- and 100,000-fold with yields of 27 +/- 15 and 19 +/- 6%, respectively. Molecular weight determination by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate and reducing agents yielded Mr of 60,500 +/- 1,800 and 65,200 +/- 400, respectively, for lipoprotein lipase and hepatic triglyceride lipase. These lipase preparations were shown to be free of detectable antithrombin by measuring its activity and by probing of Western blots of lipases with a monospecific antibody against antithrombin. In additions, probing of Western blots with concanavalin A revealed no glycoproteins corresponding to the molecular weight of antithrombin. Four stable hybridoma-producing distinct monoclonal antibodies (mAb) to hepatic triglyceride lipase were isolated. The specificity of one mAb, HL3-5, was established by its ability to immunoprecipitate hepatic triglyceride lipase catalytic activity. Interaction of HL3-5 with this lipase did not inhibit catalytic activity. The three other mAb interacted with hepatic triglyceride lipase only after denaturation of the enzyme with detergents. The relatedness of these two enzymes was examined by comparing under the same conditions the thermal inactivation, the sensitivity to sulfhydryl and reducing agents, amino acid composition, and the mobility of peptide fragments generated by cyanogen bromide cleavage. The results of these studies strongly support the view that the two enzymes are different proteins. Immunological studies confirm this conclusion. Four mAb to hepatic triglyceride lipase did not interact with lipoprotein lipase in Western blots, enzyme-linked immunosorbent assay, and immunoprecipitation experiments. These immunological studies demonstrate that several epitopes of the hepatic triglyceride lipase protein moiety are not present in the lipoprotein lipase molecule.
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PMID:Purification and characterization of human lipoprotein lipase and hepatic triglyceride lipase. Reactivity with monoclonal antibodies to hepatic triglyceride lipase. 403 Jul 67

We have investigated the effects of phospholipase A2 and C on the synthesis of prostaglandin E2 in rabbit kidney medulla and the release of fatty acids from the medulla slices. Exogenous phospholipase A2 [from Naja naja (Indian cobra) venom] and phospholipase C (from Clostridium welchii) stimulated prostaglandin E2 production in a dose-dependent manner. At the maximal effective concentrations (0.5 unit of phospholipase A2/ml, 2 units of phospholipase C/ml), phospholipase C increased prostaglandin E2 formation to the level observed with phospholipase A2. Phospholipase A2 enhanced the release only of unsaturated fatty acids, whereas phospholipase C stimulated the release of individual free fatty acids (C 16:0, C 18:0, C 18:1, C 18:2 and C 20:4). Moreover, p-bromophenacyl bromide inhibited phospholipase A2-stimulated prostaglandin E2 production and the release of fatty acids, but it had no influence on prostaglandin E2 formation and the release of fatty acids increased by phospholipase C, indicating that the stimulatory effect of phospholipase C is not mediated through the activation of endogenous phospholipase A2. These results suggest the presence of diacylglycerol lipase and monoacylglycerol lipase in the kidney and the importance of this pathway in prostaglandin synthesis by the kidney.
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PMID:Stimulation of prostaglandin E2 synthesis by exogenous phospholipase A2 and C in rabbit kidney medulla slices. 658 1

The contribution of phospholipases A2 (PLA2) and D (PLD) activation to arachidonic acid liberation and prostaglandin D2 (PGD2) formation was studied in stimulated rat peritoneal mast cells. Stimulation of the cells with ionomycin induced time-dependent and Ca(2+)-concentration-dependent increase in arachidonic acid liberation and PGD2 formation, and the Ca(2+)-dependent increase was especially remarkable at extracellular Ca2+ concentration higher than 200 microM. Staurosporine did not induce any effect on the arachidonic acid liberation, indicating that protein kinase C is not involved in the liberation. Addition of ethanol to the cells decreased the ionomycin-stimulated arachidonic acid liberation to 40% of the control, while it decreased the PGD2 formation almost completely, with the increase in phosphatidylethanol formation. Propranolol, a phosphatidate phosphohydrolase inhibitor, caused similar effects. p-Bromophenacyl bromide, a PLA2 inhibitor, inhibited partially the arachidonic acid liberation. The inhibition of the liberation by combination of p-bromophenacyl bromide and ethanol was additive and reached approximately 90%. Under the conditions used p-bromophenacyl bromide did not influence significantly the PLD activity assessed by the phosphatidylethanol formation. Histamine release was decreased by ethanol treatment to 35% of the control. These results suggest that more than half of the total arachidonic acid liberation is mediated by the sequential pathway of PLD/phosphatidate phosphohydrolase/diacylglycerol lipase and more than half of histamine release is also dependent on PLD activation, while the PGD2 formation is fully mediated by the pathway. PLA2 also contributes to arachidonic acid liberation but to a lower extent.
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PMID:Contribution of phospholipases A2 and D to arachidonic acid liberation and prostaglandin D2 formation with increase in intracellular Ca2+ concentration in rat peritoneal mast cells. 750 86

Five microbial lipases from Chromobacterium viscosum, Candida cylindracea, Pseudomonas (source Fluka), Pseudomonas (source Genzyme) and lipoprotein lipase ex Microbial (Genzyme) have been screened for lactonisation activity towards 16-hydroxyhexadecanoic acid (HHA) in a variety of different w/o microemulsion systems. With the exception of Candida cylindracea (CC), all the lipases exhibited lactonisation activity although they were inherently more active in microemulsion systems based on the anionic surfactant sodium bis(2-ethylhexyl)sulphosuccinate (AOT) than in those based on the cationic surfactant cetyltrimethylammonium bromide (CTAB). Lactone yields are typically 50-60% and are markedly better than those reported previously using microemulsions in combination with chemical catalysts. Lipase stability is superior in the CTAB microemulsion systems, while lipase stability in the low water content AOT microemulsion systems was still good with the exception of CC lipase, which is rapidly inactivated. Buffering the water pools of AOT microemulsions using diglycine buffer at pH 8.0 improved biocatalyst stability. The lactonisation activity of lipases in CTAB w/o microemulsion systems compares favourably with that obtained using the same preparations as a solid suspension in the corresponding water-saturated organic solvent. In addition, the unusual solubility properties of microemulsions allowed the use of considerably higher concentrations of substrate in the microemulsion systems as compared to water-saturated organic solvents such as n-heptane. Lactone yields obtained at equivalent concentrations in the corresponding organic solvents containing conventional condensation catalysts were consistently measured at approx. 10%.
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PMID:Macrocyclic lactone synthesis by lipases in water-in-oil microemulsions. 754 59

We have developed a novel one-step pool screening PCR procedure which is based on the principles of amplification refractory mutation system (ARMS) and competitive oligonuleotide priming (COP) PCR. In addition to the usual primers, this approach uses two allele-specific competitive oligonucleotides, one of which is 3'-end labeled with a dideoxynucleotide and blocks amplification of the wild-type allele. An allele-specific product is generated only in the presence of the mutation. The introduction of an allele-specific competitive blocker oligonucleotide improves the specificity and robustness of ARMS-PCR. Further its sensitivity is dramatically increased, which allows detection of one mutant allele in a large excess of wild-type-bearing genomic DNA by electrophoresis in an ethidium bromide-stained agarose gel (up to 1 in 10(4) alleles). This makes the method ideal for nonradioactive pool screening. The successful application of the method has been demonstrated for four different point mutations, two in the apolipoprotein B gene (R3500Q, R3531C) which result in familial defective apolipoprotein B-100, one in the CFTR gene (R1162X), and one in the gene for lipoprotein lipase (G188E).
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PMID:Allele-specific competitive blocker PCR: a one-step method with applicability to pool screening. 758


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