Lipoprotein lipase immobilized

The liver synthesizes two enzymes involved in intra-plasmic lipid metabolism hepatic triglyceride lipase (HTL) and lecithin-cholesterol-acyltransferase (LCAT). The liver is further involved in the modification of circulatory lipoproteins as the site of synthesis for cholesterol-ester transfer protein (CETP). Free fatty acids are in general potentially toxic to the liver cell. Therefore they are immobilized by being bound to the intrinsic hepatic fatty acid-binding protein (hFABP) in the cytosol. The activity of this protein is stimulated by oestrogens and inhibited by testosterone. Peripheral lipoprotein lipase (LPL), which is required for the regulation of lipid metabolism, is synthesized in the endothelial cells (mainly in the fatty tissue and musculature). 

Immobilized enzymes are currently the object of considerable interest. This is due to the expected benefits over soluble enzymes or alternative technologies. The number of applications of immobilized enzymes is increasing steadily [5]. Occasionally, however, experimental investigations have produced unexpected results such as a significant reduction or even an increase in activity compared with soluble enzymes. Thus, cross-linked crystals of subtilisin showed 27 times less activity in the aqueous hydrolysis of an amino acid ester compared to equal amounts of soluble enzyme [6]. On the other hand, in the application of lipoprotein lipase in the solvent-mediated synthesis of esters there was a 40-fold increase in activity using immobilized or otherwise modified enzyme preparations as compared to enzyme powder [7]. 

Thus, triglycerides have been determined with lipoprotein lipase (EC 3.1.1.34) immobilized on CPG with a pore size of 2000 A. The assay buffer was 0.1 M Tris buffer, pH 8.0, containing 0.5% Triton X-100. The linear response was 0.05-10 mM for tributyrin and 0.1-5 mM for triolein [24]. 

Functions.—Heparin fractionated by gel filtration appeared to bind to two sites on antithrombin III (association constants 0.6 x 10 and 0.2 x 10 moll" ), whereas heparin prepared by affinity chromatography on matrix-bound antithrombin III appeared to bind to only one site (association constant 2.3 x 10 moll ). These results suggest that one of the binding sites on antithrombin III does not bind the most active heparin components, but accommodates heparin-like molecules which, although similar in size to the active heparin components, have little or no anticoagulant activity. Heparins with high or low affinities for antithrombin III exhibited no differences in their abilities to bind lipoprotein lipase. Studies of the interaction between the lipoprotein lipase from cow s milk and Sepharose-immobilized heparin have shown that heparin is poly-disperse. Whereas heparin facilitated complex formation between a-thrombin and antithrombin III, it had little effect on the interaction between p-thrombin and antithrombin III. ... 

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