Pancreatic porcine phospholipase A2 catalyzed hydrolysis of phosphatidylcholine in lecithin-bile salt mixed micelles: kinetic studies in a lecithin-sodium cholate system.
Pancreatic porcine phospholipase A2 catalyzed hydrolysis of phosphatidylcholine in bile salt lecithin mixed micelles has been studied, utilizing a series of assay mixtures for which the micellar size, weight, and composition had been experimentally determined. Under these conditions the enzymatic hydrolysis is dependent on the phosphatidylcholine-to-sodium cholate molar ratio within the mixed micelle rather than the bulk concentration of the phospholipid in the mixture: at 5 mM phosphatidylcholine, variation of the NPC/NNaCh ratio from 0.2 to 2.0 increases the enzymatic activity from 82 to 933 mumol/min/mg protein. The initial rates are linear throughout the entire series of assay mixtures, the activity vs micellar concentration curves exhibit saturation behavior, and treatment of the data according to the "surface-as-cofactor" theory provides linear double-reciprocal plots which intersect in one point. The assay system should be applicable for detailed kinetic studies of lipolytic enzymes, including mammalian phospholipases which exhibit rather low activities toward lecithin-Triton X-100 mixed micelles. The system should also provide a convenient basis for mechanistic studies involving the use of inhibitory phospholipid substrate analogs.
D000818:Animals; D001647:Bile Acids and Salts / Q000378:metabolism*; D002384:Catalysis; D019826:Cholic Acid; D002793:Cholic Acids; D004546:Elapid Venoms / Q000032:analysis; D006868:Hydrolysis; D007700:Kinetics; D008823:Micelles; D010179:Pancreas / Q000201:enzymology*; D010713:Phosphatidylcholines / Q000378:metabolism*; D010740:Phospholipases / Q000502:physiology*; D010741:Phospholipases A / Q000502:physiology*; D054467:Phospholipases A2; D013552:Swine
D001647:Bile Acids and Salts; D002793:Cholic Acids; D004546:Elapid Venoms; D008823:Micelles; D010713:Phosphatidylcholines; D010740:Phospholipases; D010741:Phospholipases A; D054467:Phospholipases A2; D019826:Cholic Acid