Hydrolysis of emulsified mixtures of triacylglycerols by pancreatic lipase. (1/105)

Hydrolysis of the emulsified mixture of short-chain triacylglycerols by porcine pancreatic lipase in the presence of procolipase and micellar sodium taurodeoxycholate has been studied. Increase in the content of tributyrin and trioctanoin in the mixture with triacetin had highly cooperative effects on the formation of the interfacial lipase procolipase complex. Abrupt enhancement of the complex stability was observed in the presence of 0.4-0.6 mol mol-1 of tributyrin or 0.58 mol mol-1 of trioctanoin in the substrate phase. The affinity of lipase towards interfacially bound procolipase for the trioctanoin containing 0.07-0.42 mol mol-1 of triacetin was approximately three times higher than that for pure trioctanoin. The cooperative processes involved in complex formation did not contribute to the affinity of the interfacial lipase/(pro)colipase complex towards substrate molecules and its catalytic activity.  (+info)

Pancreatic lipase/colipase-mediated triacylglycerol hydrolysis is required for cholesterol transport from lipid emulsions to intestinal cells. (2/105)

This study tested the hypothesis that dietary cholesterol uptake by intestinal cells is dependent on the structure and composition of the lipid carriers in the extracellular milieu. In in vivo experiments with female C57BL/6 mice, cholesterol absorption from phospholipid/triacylglycerol emulsions was significantly reduced by administration of tetrahydrolipstatin, an inhibitor of pancreatic lipase. This inhibitor had no effect on the absorption of cholesterol from phospholipid vesicles. The importance of pancreatic-lipase-mediated triacylglycerol hydrolysis for cholesterol transport from emulsions to intestinal cells was confirmed by in vitro experiments with rat IEC-6 intestinal cells. Cellular uptake of cholesterol from emulsions with a phospholipid/triacylglycerol molar ratio of <0.3 could be stimulated by pancreatic lipase/colipase hydrolysis of the core neutral lipids. However, pancreatic lipase/colipase was ineffective in hydrolysing triacylglycerols in emulsions with a phospholipid/triacylglycerol molar ratio of >0.3. Phospholipase A2-mediated hydrolysis of the surface phospholipids was necessary prior to triacylglycerol hydrolysis in these phospholipid-rich emulsions and to the stimulation of cholesterol transport from these particles to IEC-6 cells. The data also revealed that minimal triacylglycerol hydrolysis was sufficient to significantly increase cholesterol transport from lipid emulsions to the intestinal cells. Thus the products of triacylglycerol hydrolysis, namely monoacylglycerol and non-esterified fatty acids, are key determinants in mediating cholesterol transport from lipid emulsions to intestinal cells. Taken together, these results support the hypothesis that remodelling of the surface and core components of lipid carriers is necessary prior to absorption of dietary cholesterol from the gastrointestinal tract.  (+info)

Colipase: structure and interaction with pancreatic lipase. (3/105)

Colipase is a small protein cofactor needed by pancreatic lipase for the efficient dietary lipid hydrolysis. It binds to the C-terminal, non-catalytic domain of lipase, thereby stabilising an active conformation and considerably increasing the overall hydrophobic binding site. Structural studies of the complex and of colipase alone have clearly revealed the functionality of its architecture. Interestingly, a structural analogy has recently been discovered between colipase and a domain in a developmental protein (Dickkopf), based on sequence analogy and homology modeling. Whether this structural analogy implies a common function (lipid interaction) remains to be clarified. Structural analogies have also been recognised between the pancreatic lipase C-terminal domain, the N-terminal domains of lipoxygenases and the C-terminal domain of alpha-toxin. These non-catalytic domains in the latter enzymes are important for interaction with membranes. It has not been established if these domains are also involved in eventual protein cofactor binding as is the case for pancreatic lipase.  (+info)

Critical role of micelles in pancreatic lipase activation revealed by small angle neutron scattering. (4/105)

In the duodenum, pancreatic lipase (PL) develops its activity on triglycerides by binding to the bile-emulsified oil droplets in the presence of its protein cofactor pancreatic colipase (PC). The neutron crystal structure of a PC-PL-micelle complex (Hermoso, J., Pignol, D., Penel, S., Roth, M., Chapus, C., and Fontecilla-Camps, J. C. (1997) EMBO J. 16, 5531-5536) has suggested that the stabilization of the enzyme in its active conformation and its adsorption to the emulsified oil droplets are mediated by a preformed lipase-colipase-micelle complex. Here, we correlate the ability of different amphypathic compounds to activate PL, with their association with PC-PL in solution. The method of small angle neutron scattering with D(2)O/H(2)O contrast variation was used to characterize a solution containing PC-PL complex and taurodeoxycholate micelles. The resulting radius of gyration (56 A) and the match point of the solution indicate the formation of a ternary complex that is similar to the one observed in the neutron crystal structure. In addition, we show that either bile salts, lysophospholipids, or nonionic detergents that form micelles with radii of gyration ranging from 13 to 26 A are able to bind to the PC-PL complex, whereas smaller micelles or nonmicellar compounds are not. This further supports the notion of a micelle size-dependent affinity process for lipase activation in vivo.  (+info)

Effect of high-fat diet, surrounding temperature, and enterostatin on uncoupling protein gene expression. (5/105)

Nonshivering thermogenesis induced in brown adipose tissue (BAT) during high-fat feeding is mediated through uncoupling protein 1 (UCP1). UCP2 is a recently identified homologue found in many tissues. To determine the role of UCP1 and UCP2 in thermoregulation and energy balance, we investigated the long-term effect of high-fat feeding on mRNA levels in mice at two different ambient temperatures. We also treated mice with the anorectic peptide enterostatin and compared mRNA levels in BAT, white adipose tissue (WAT), stomach, and duodenum. Here, we report that high-fat feeding at 23 degrees C increased UCP1 and UCP2 levels in BAT four- and threefold, respectively, and increased UCP2 levels fourfold in WAT. However, at 29 degrees C, UCP1 decreased, whereas UCP2 remained unchanged in BAT and increased twofold in WAT. Enterostatin increased UCP1 and decreased UCP2 mRNA in BAT. In stomach and duodenum, high-fat feeding decreased UCP2 mRNA, whereas enterostatin increased it. Our results suggest that the regulation of uncoupling protein mRNA levels by high-fat feeding is dependent on ambient temperature and that enterostatin is able to modulate it.  (+info)

Fats infused intraduodenally affect the postprandial secretion of the exocrine pancreas and the plasma concentration of cholecystokinin but not of peptide YY in growing pigs. (6/105)

In pigs, the spontaneous secretion of the exocrine pancreas and the release of cholecystokinin (CCK) and peptide YY (PYY) after intraduodenal infusion of fully saturated synthetic fats differing in chain length was studied. Growing pigs (n = 6) were prepared with pancreatic duct catheters, duodenal T-cannulas and catheters placed in the jugular vein. The pigs were fed 2 g/100 g body twice daily. Beginning with the morning feeding, a medium-chain triglyceride (MCT: glycerol tricaprylate), a long-chain triglyceride (LCT: glycerol tristearate) or saline was infused at a rate of 0.1 g/100 g body. Pancreatic juice was collected, beginning 1 h preprandially until 3 h postprandially. Blood samples were obtained 15 min preprandially and 15, 45, 90 and 150 min postprandially. The infusion of MCT evoked a change in the trend of the curve for the volume of secretion of pancreatic juice, lipase and colipase concentrations and outputs. The trend of the curve did not change over time for CCK and PYY. Differences between the trends of the curves for the saline and MCT treatment were observed for volume of secretion, protein output, lipase content and output, trypsin and colipase output. Differences in the trends of the curves between MCT and LCT were obtained for the outputs of protein, lipase and colipase. Plasma CCK levels were lower as a result of the MCT treatment compared with the saline and LCT treatments. The results suggest an immediate, distinguished response of the porcine exocrine pancreas to fats differing in chain length.  (+info)

Interactions of bile salt micelles and colipase studied through intermolecular nOes. (7/105)

Colipase is a small protein (10 kDa), which acts as a protein cofactor for the pancreatic lipase. Various models of the activated ternary complex (lipase-colipase-bile salt micelles) have been proposed using detergent micelles, but no structural information has been established with bile salt micelles. We have investigated the organization of sodium taurodeoxycholate (NaTDC) micelles and their interactions with pig and horse colipases by homonuclear nuclear magnetic resonance (NMR) spectroscopy. The NMR data supply evidence that the folding of horse colipase is similar to that already described for pig colipase. Intermolecular nuclear Overhauser effects have shown that two conserved aromatic residues interact with NaTDC micelles.  (+info)

The lipase C-terminal domain. A novel unusual inhibitor of pancreatic lipase activity. (8/105)

In vertebrates, dietary fat digestion mainly results from the combined effect of pancreatic lipase, colipase, and bile. It has been proposed that in vivo lipase adsorption on oil-water emulsion is mediated by a preformed lipase-colipase-mixed micelle complex. The main lipase-colipase binding site is located on the C-terminal domain of the enzyme. We report here that in vitro the isolated C-terminal domain behaves as a potent noncovalent inhibitor of lipase and that the inhibitory effect is triggered by the presence of micelles. Lipase inhibition results from the formation of a nonproductive C-terminal domain-colipase-micelle ternary complex, which competes for colipase with the active lipase-colipase-micelle ternary complex, thus diverting colipase from its lipase-anchoring function. The formation of such a complex has been evidenced by molecular sieving experiments. This nonproductive complex lowers the amount of active lipase thus reducing lipolysis. Preliminary experiments performed in rats show that the C-terminal domain also behaves as an inhibitor in vivo and thus could be considered a potential new tool for specifically reducing intestinal lipolysis.  (+info)

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