Characterization of a bile salt-dependent cholesteryl ester hydrolase activity secreted from HepG2 cells

Winkler, Karl; Harrison, Earl H.; Marsh, Julian B.; Glick, Jane M.; Ross, Alastair

doi:10.1016/0005-2760(92)90285-4

Cited by 45 publications

(33 citation statements)

References 27 publications

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“…BSDL is synthesized to a limited extent by human-monocytes-macrophages (32), endothelial cells (33,34), eosinophils (35), and the liver (22,36,37). BSDL has also been localized in necrotic areas of the pancreas, consecutive to an acute pancreatitis (38).…”

Section: Discussionmentioning

confidence: 99%

Site-directed Mutagenesis of the Basic N-terminal Cluster of Pancreatic Bile Salt-dependent Lipase

Aubert¹,

Sbarra

Petit-Thévenin

et al. 2002

Journal of Biological Chemistry

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Previous studies have postulated the presence of a heparin-binding site on the bile salt-dependent lipase (BSDL), whereas two bile salt-binding sites regulate the enzyme activity. One of these sites may overlap with the tentative heparin-binding site at the level of an N-terminal basic cluster consisting of positive residues Lys 32 , Lys 56 , Lys 61 , Lys 62 , and Arg 63 . The present study uses specific site-directed mutagenesis to determine the functional significance of this basic cluster. Mutations in this sequence resulted in recombinant enzymes that were able to bind to immobilized and to cell-associated heparin before moving throughout intestinal cells. Recombinant BSDL was fully active on soluble substrate, but mutants were less active on micellar cholesteryl oleate in comparison with the wild-type enzyme. Activation studies by primary (sodium taurocholate) and by secondary (sodium taurodeoxycholate) bile salts revealed that the activation of BSDL by sodium taurocholate at concentrations below the critical micellar concentration, and not that evoked by micellar bile salts, was affected by substitutions, suggesting that this Nterminal basic cluster likely represents the specific bile salt-binding site of BSDL. Substitutions also affected the activation of the enzyme promoted by anionic phospholipids, extending the function of this site to that of a cationic regulatory site susceptible to accommodate anionic ligands.

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Section: Discussionmentioning

confidence: 99%

Site-directed Mutagenesis of the Basic N-terminal Cluster of Pancreatic Bile Salt-dependent Lipase

Aubert¹,

Sbarra

Petit-Thévenin

et al. 2002

Journal of Biological Chemistry

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“…The hepatic CEL was found to be present in specific endosomal compartments of hepatocytes (127). In a separate study, Winkler et al (128) showed that the hepatic-derived CEL can be secreted directly by liver cells. Thus, the CEL found in the endosomal compartment of the liver may be recaptured enzyme secreted by hepatic cells.…”

Section: Hepatic Lipoprotein Metabolismmentioning

confidence: 98%

Carboxyl ester lipase

Hui

Howles

2002

Journal of Lipid Research

196

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Carboxyl ester lipase (CEL), previously named cholesterol esterase or bile salt-stimulated (or dependent) lipase, is a lipolytic enzyme capable of hydrolyzing cholesteryl esters, tri-, di-, and mono-acylglycerols, phospholipids, lysophospholipids, and ceramide. The active site catalytic triad of serine-histidine-aspartate is centrally located within the enzyme structure and is partially covered by a surface loop. The carboxyl terminus of the protein regulates enzymatic activity by forming hydrogen bonds with the surface loop to partially shield the active site. Bile salt binding to the loop domain frees the active site for accessibility by water-insoluble substrates. CEL is synthesized primarily in the pancreas and lactating mammary gland, but the enzyme is also expressed in liver, macrophages, and in the vessel wall. In the gastrointestinal tract, CEL serves as a compensatory protein to other lipolytic enzymes for complete digestion and absorption of lipid nutrients. Importantly, CEL also participates in chylomicron assembly and secretion, in a mechanism mediated through its ceramide hydrolytic activity. Cell culture studies suggest a role for CEL in lipoprotein metabolism and oxidized LDL-induced atherosclerosis. Thus, this enzyme, which has a wide substrate reactivity and diffuse anatomic distribution, may have multiple functions in lipid and lipoprotein metabolism, and atherosclerosis. Carboxyl ester lipase (CEL), previously named pancreatic cholesterol esterase and also known as bile salt-stimulated (or -dependent) lipase, is a nonspecific lipolytic enzyme capable of hydrolyzing cholesteryl esters, tri-, di-, and mono-acylglycerols, phospholipids, lysophospholipids, and ceramide (1-3). The hydrolysis of water-insoluble carboxyl esters with long chain fatty acyl groups by CEL requires its activation by bile salt. However, CEL hydrolysis of water-soluble substrates such as carboxyl esters with short chain fatty acids or lysophospholipids does not have an absolute dependence on bile salt activation. The CEL protein is synthesized primarily in the pancreatic acinar cells and lactating mammary glands of higher mammals. Small amounts of CEL are found in other tissues, particularly the liver, macrophages, endothelial cells, and eosinophils (4-10). Extensive research has been performed in recent years on the structure-function relationship of this protein as well as its physiological role in lipid metabolism. A significant body of work has also been devoted to studying CEL processing and transport in pancreatic acinar cells. The latter topic was reviewed recently (11) and will be discussed only briefly in this article when appropriate. This article will present an updated review on the structure-function studies of CEL as well as an in-depth discussion on the current understanding of its role in lipid metabolism. PROTEIN STRUCTURE-FUNCTION RELATIONSHIP Active siteThe nucleotide sequence of CEL from various species shows that it is a highly conserved protein belonging to the ␣ / ␤ hydrolase family. The carb...

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“…For example, there are reports of CEL activity and/or mRNA in rat liver (e.g., references 5-7), human breast (8), human hepatocarcinoma HepG2 cells (9), human placenta (10), rat (11) and rabbit heart (12), and rat and rabbit aorta (13,14). Particularly intriguing is the evidence that CEL is found in the blood plasma of a number of mammals, including humans (15), rats (11), dogs, and goats (16).…”

Section: Introductionmentioning

confidence: 99%

Pancreatic carboxyl ester lipase: a circulating enzyme that modifies normal and oxidized lipoproteins in vitro.

Shamir¹,

Johnson²,

Morlock-Fitzpatrick³

et al. 1996

J. Clin. Invest.

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Pancreatic carboxyl ester lipase (CEL) hydrolyzes cholesteryl esters (CE), triglycerides (TG), and lysophospholipids, with CE and TG hydrolysis stimulated by cholate. Originally thought to be confined to the gastrointestinal system, CEL has been reported in the plasma of humans and other mammals, implying its potential in vivo to modify lipids associated with LDL, HDL (CE, TG), and oxidized LDL (lysophosphatidylcholine, lysoPC). We measured the concentration of CEL in human plasma as 1.2 Ϯ 0.5 ng/ml (in the range reported for lipoprotein lipase). Human LDL and HDL 3 reconstituted with radiolabeled lipids were incubated with purified porcine CEL without or with cholate (10 or 100 M, concentrations achievable in systemic or portal plasma, respectively). Using a saturating concentration of lipoprotein-associated CE (4 M), with increasing cholate concentration there was an increase in the hydrolysis of LDL-and HDL 3 -CE; at 100 M cholate, the percent hydrolysis per hour was 32 Ϯ 2 and 1.6 Ϯ 0.1, respectively, indicating that CEL interaction varied with lipoprotein class. HDL 3 -TG hydrolysis was also observed, but was only ‫ف‬ 5-10% of that for HDL 3 -CE at either 10 or 100 M cholate. Oxidized LDL (OxLDL) is enriched with lysoPC, a proatherogenic compound. After a 4-h incubation with CEL, the lysoPC content of OxLDL was depleted 57%. Colocalization of CEL in the vicinity of OxLDL formation was supported by demonstrating in human aortic homogenate a cholate-stimulated cholesteryl ester hydrolytic activity inhibited by anti-human CEL IgG. We conclude that CEL has the capability to modify normal human LDL and HDL composition and structure and to reduce the atherogenicity of OxLDL by decreasing its lysoPC content.

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Characterization of a bile salt-dependent cholesteryl ester hydrolase activity secreted from HepG2 cells

Cited by 45 publications

References 27 publications

Site-directed Mutagenesis of the Basic N-terminal Cluster of Pancreatic Bile Salt-dependent Lipase

Site-directed Mutagenesis of the Basic N-terminal Cluster of Pancreatic Bile Salt-dependent Lipase

Carboxyl ester lipase

Pancreatic carboxyl ester lipase: a circulating enzyme that modifies normal and oxidized lipoproteins in vitro.

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