Purification and characterization of rat adipose tissue lipoprotein lipase.
Lipoprotein lipase (EC 3.1.1.34) extracted from adipose tissue of glucose-fed rats with 5 mM-sodium barbital, pH 7.5, containing 20% (v/v) glycerol and 0.1% (v/v) Triton X-100, was partially purified by affinity chromatography on heparin linked to Sepharose 4B. Sodium dodecyl sulphate/polyacrylamide-gel electrophoresis of the partially purified enzyme preparation revealed the presence of two major Coomassie-staining bands (mol.wts. 62 000 and 56 000) as well as a number of minor bands. Treatment of partially purified enzyme with [1,3-3H]di-isopropyl fluorophosphate resulted in the incorporation of radiolabel into the band of mol.wt. 56 000, but not into the band of mol.wt. 62 000. Both the amount of the 56 000-mol.wt. polypeptide and the incorporation of [1,3-3H]di-isopropyl fluorophosphate into this band were greatly reduced in the enzyme preparations isolated from adipose tissue of 48 h-starved rats. whereas the amount of the 62 000-mol.wt. polypeptide was unaffected by starvation. Purification of lipoprotein lipase from adipose tissue of glucose-fed rats was also carried out using affinity chromatography on Sepharose 4B linked to heparin with low affinity for antithrombin-III. This procedure resulted in the presence of a single band of mol.wt. 56 000 on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. These results suggest that the polypeptide of mol.wt. 56 000 corresponds to the subunit of lipoprotein lipase, whereas the 62 000-mol.wt. polypeptide probably represents antithrombin-III.
D000273:Adipose Tissue / Q000201:enzymology*; D000818:Animals; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D006493:Heparin; D007531:Isoflurophate / Q000494:pharmacology; D008071:Lipoprotein Lipase / Q000037:antagonists & inhibitors / Q000302:isolation & purification*; D008297:Male; D008970:Molecular Weight; D051381:Rats; D011919:Rats, Inbred Strains; D012965:Sodium Chloride; D013217:Starvation / Q000201:enzymology
D007531:Isoflurophate; D012965:Sodium Chloride; D006493:Heparin; D008071:Lipoprotein Lipase