Lipoprotein lipase enzyme, activity

Lipase was first isolated from skim milk and characterized by Fox and Tarassuk in 1967. The enzyme was optimally active at pH 9.2 and 37°C and found to be a serine enzyme (inactivated by organophosphates). A lipoprotein lipase (LPL activated by lipoprotein co-factors) was demonstrated in milk by Korn in 1962 and was isolated by Egelrud and Olivecrona in 1972. LPL is, in fact, the principal indigenous lipase in milk and most recent work has been focused accordingly. The molecule has been characterized at the molecular, genetic, enzymatic and physiological levels (see Olivecrona et al, 1992). 

The use of vacutainer tubes and heparin was shown to alter the determination of protein binding. Heparin was shown to decrease the plasma binding of certain drugs including phenytoin, propranolol, lidocaine, diazepam, quinidine, and verapamil. This is also an in vitro artifact attributable to continued ex vivo activity of the lipoprotein lipase enzyme and accumulation of fatty acids in the blood collection tube. 

Lipoprotein lipase is an enzyme found in the capillaries that catalyzes the hydrolysis of triacylglycerols in chylomicrons to glycerol and fatty acids (Figure 18.6, Figure 18.7). Lipoprotein lipase is activated by apoprotein C-II, which is found in all of the lipoprotein complexes except LDLs. Apoprotein C-1 may also be involved in activation of lipoprotein lipase. 

Fig. 32.13. Fate of chylomicrons. Chylomicrons are synthesized in intestinal epithelial cells, secreted into the lymph, pass into the blood, and become mature chylomicrons (see Fig. 32.11). On capillary walls in adipose tissue and muscle, lipoprotein lipase (LPL) activated by ApoCn digests the triacylglycerols (TG) of chylomicrons to fatty acids and glycerol. Fatty acids (FA) are oxidized in muscle or stored in adipose cells as triacylglycerols. The remnants of the chylomicrons are taken up by the liver by receptor-mediated endocytosis. Lysosomal enzymes within the hepatocyte digest the remnants, releasing the products into the cytosol.

The enzyme responsible for lipolytic release of fatty acids from adipose tissue triglycerides is most likely not lipoprotein lipase. The activity of this enzyme increases under conditions of maximum uptake but decreases when maximum release is induced (Cherkes and Gordon 1959 Salaman and Robinson 1961). Any function of lipoprotein lipase in the transport of fat would therefore be in the uptake and not in the release. 

Lipoprotein lipase is activated by high levels of insulin. It acts to extract FAs and glycerol from chylomicrons in the circulation, which are taken up by the adipocytes and re-esterified into TAG to be stored. Insulin inactivates hormone-sensitive lipase to ensure that TAG is not degraded. (Be careful not to get the two enzymes confused.) In the fed state the brain and erythrocytes exclusively use glucose as their fuel supply. Muscle uses glucose as its main fuel, and is also able to metabolize branched amino acids (leucine, isoleucine and valine) as well. 

Gene activated Lipoprotein lipase fatty acid transporter protein adipocyte fatty acid binding protein acyl-CoA synthetase malic enzyme GLUT-4 glucose transporter phosphoenolpyruvate carboxykinase... 

Sample Collection and Enzyme Stability. Serum samples are collected with chemically clean, sterile glassware. Blood is allowed to clot at room temperature, the clot is gently separated from the test tube with an applicator stick, and the blood is centrifuged for 10 minutes at 1,000 g. If the red cells are known to contain the enzymes whose activity is being measured, as in the case of LD, even slightly hemolyzed serums must be discarded. When acid phosphatase is to be measured, the serum should be placed immediately in ice and processed as soon as possible, or it should be acidified by the addition of a small amount of sodium citrate. Anticoagulants such as EDTA, fluoride and oxalate inhibit some serum enzymes. However, heparin activates serum lipoprotein lipase. 

Diagnosis of lipoprotein lipase deficiency is based on low or absent enzyme activity with normal human plasma or apolipoprotein C-II, a cofactor of the enzyme. 

Lipoprotein (LPLase) is required for the metabolism of both chylomicrons and VLDL. This enzyme is induced by insulin and transported to the luminal surface of capillary endothelium where it is in direct contact with the blood. Lipoprotein lipase hydrolyzes the fiitty adds from triglycerides carried by ch)4oinicrons and VLDL and is activated by apoC-II. 

The best-known effect of APOE is the regulation of lipid metabolism (see Fig. 10.13). APOE is a constituent of TG-rich chylomicrons, VLDL particles and their remnants, and a subclass of HDL. In addition to its role in the transport of cholesterol and the metabolism of lipoprotein particles, APOE can be involved in many other physiological and pathological processes, including immunoregu-lation, nerve regeneration, activation of lipolytic enzymes (hepatic lipase, lipoprotein lipase, lecithin cholesterol acyltransferase), ligand for several cell receptors, neuronal homeostasis, and tissue repair (488,490). APOE is essential... 

During lactation, the enzyme is active in mammary gland and the released fatty acids are taken up and used to synthesise triacylglycerol for the milk. A few days before parturition, in preparation for lactation, the activity of lipoprotein lipase is increased in the mammary gland. 

Lipoprotein lipase, an enzyme responsible for the hydrolysis of the triglyceride moiety of lipoproteins, has been shown to be activated by serum high-density lipoproteins (B4, K5, S6, S14), by very low density... 

Current available information does not permit definitive conclusions on the nature, specificity, and mechanism of action of the protein cofactor (s) of lipoprotein lipase. It is verj difiicult to correlate the observations described above (summarized in Table 10) since the enzyme preparations used were not pure or well characterized, and were derived from various sources. For instance, two species of lipoprotein lipase have been reported to exist in rat adipose tissue (G4), and major differences between enzymes of liver and adipose tissue have been noted (G16). Also, the nature of the apoprotein preparations employed as protein cofactor (s) of lipoprotein lipase has not been clearly specified in all the studies contaminated materials may account for the spurious results observed. At present, it is not known how apoproteins such as apo Glu, apo Ala, and apo Ser could exhibit their activator or inhibitor activity on lipoprotein lipase. If these different apoproteins indeed prove to be cofactors for lipoprotein lipase, the nature of the lipid-protein specificity must be established and thus the role played by carbohydrates, since some of these apoproteins are glycoproteins. 

P3. Posner, I., and Morales, A., Mechanisms of enzyme and substrate activation by lipoprotein lipase cofactors. I. A specific requirement of physiological concentrations of calcium for enzyme activity. J. Biol. Chem. 247, 2255-2265 (1972). 

D. Lipoprotein lipase. Fenohbrate is a hypotriglyc-eridemic drug that lowers plasma triglycerides by increasing the activity of hpoprotein lipase, the enzyme responsible for disassembly of triglycerides in serum lipoproteins (VLDL, IDL and chylomicrons). 

These activate lipoprotein lipase which is a key enzyme in degradation of VLDL resulting in lower circulating triglycerides. These drugs lower triglyceride levels by 20-50% with 10-15% decrease in LDL cholesterol and a 10-15% increase in HDL cholesterol. 

In the capillaries of these tissues, the extracellular enzyme lipoprotein lipase, activated by apoC-II, hydrolyzes triacylglycerols to fatty acids and glycerol (step... 

Modification of nascent chylomicron particles The particle released by the intestinal mucosal cell is called a "nascent" chylomicron because it is functionally incomplete. When it reaches the plasma, the particle is rapidly modified, receiving apo E (which is recognized by hepatic receptors) and C apolipoproteins, The latter include apo C-ll, which is necessary for the activation of lipoprotein lipase, the enzyme that degrades the triacylglycerol contained in the chylomicron (see below). The source of these apolipoproteins is circulating HDL (see Figure 18.16). 

Downey (1980) reasoned that although milk lipoprotein lipase is present in sufficient amounts to cause extensive hydrolysis and potential marked flavor impairment, this does not happen in practice for the following reasons (1) the fat globule membrane separates the milk fat from the enzyme, whose activity is further diminished by (2) its occlusion by casein micelles (Downey and Murphy 1975) and by (3) the possible presence in milk of inhibitors of lipolysis (Deeth and Fitz-Gerald 1975). The presence in milk of activators and their relative concentration may also determine whether milk will be spontaneously rancid or not (Jellema 1975 Driessen and Stadhouders 1974A Murphy et al. 1979 Anderson 1979). 

Anderson, M. 1979. Enzyme immunoassay for measuring lipoprotein lipase activator in milk. J. Dairy Sci. 62, 1380-1383. 

Pasteurization inactivates many enzymes, including alkaline phosphatase and lipoprotein lipase. The absence of active alkaline phosphatase in cheese is often used to determine if the milk has been properly pasteurized prior to cheesemaking. Since pasteurization kills most of the lactic acid bacteria in milk, the lactic acid developed during cheese-... 

Both chylomicrons and VLDL particles undergo similar processes in the capillary blood vessels, where their triacylglycerols are hydrolyzed to glycerol and free fatty acids by lipoprotein lipase 40 42a This enzyme requires for its activity the apolipoprotein C-II which is present in the chylomicrons and VLDL particles. Lipoprotein lipase is also known as the "clearing... 

Chylomicrons transport dietary triacylglycerol and cholesteryl ester from the intestine to other tissues in the body. Very-low-density lipoprotein functions in a manner similar to the transport of endogenously made lipid from the liver to other tissues. These two types of triacylglycerol-rich particles are initially degraded by the action of lipoprotein lipase, an extracellular enzyme that is most active within the capillaries of adipose tissue, cardiac and skeletal muscle, and the lactating mammary gland. Lipoprotein lipase catalyzes the hydrolysis of triacylglycerols (see fig. 18.3). The enzyme is specifically activated by apoprotein C-II, which... 

Apolipoprotein C-II can also be isolated from VLDL or HDL (H20, L5, N3). It contains 78 residues (J3) and has been shown by Chou-Fasman analysis to bind phospholipids (M26, M40), with three predicted helical sequences (M26). ApoC-II has attracted a great deal of attention because it activates one of the most important enzymes in plasma lipid metabolism, lipoprotein lipase, responsible for the hydrolysis of triglyceride in chylomicrons and VLDL. Sparrow and Gotto have summarized a number of studies on structure-function relationships (S52). These, taken together, indicate that there are separate functional domains in apoC-II, in that lipoprotein lipase activation is mediated by residues 55-78 and phospholipid binding by... 

Chylomicrons are triglyceride rich and contain apolipoprotein B-48 and the A types. The latter are synthesized in the intestinal tract cells. Additional apoproteins are transferred to the chylomicrons from HDL in circulation the apoE and apoC types. Their site of synthesis is the liver. The chylomicrons are subject to degradation by lipoprotein lipase in the peripheral tissue, especially adipose tissue. Lipoprotein lipase activity is increased by increased blood insulin levels. This enzyme is extracellular, attached to the capillary endothelial cells, and activated by ApoC-II, which is present in the chylomicrons. Lipoprotein lipase causes the hydrolysis of triglycerides, thus decreasing chylomicron size... 

Situations in which the blood insulin/glucagon ratio is higher than normal lead to fatty acid and cholesterol biosynthesis, whereas low insulin/glucagon ratios are characterized by lipolysis, increased activity of the /3-oxidation pathway, and a low level of cholesterol biosynthetic activity. Enzymes that are either activated by insulin or derepressed by low glucagon levels are lipoprotein lipase, which... 

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