Kinetics of hydrolysis of dispersions of saturated phosphatidylcholines by Crotalus atrox phospholipase A2.
Dispersions of lamellar phase dipalmitoyl phosphatidylcholine (DDPC) and dimyristoyl phosphatidylcholine (DMPC) in 0.01 M CaCl2 were subjected to hydrolysis by phospholipase A2 (EC 3.1.1.4) from Crotalus atrox venom. The reaction was followed continuously by titrating the released fatty acids. For hydrolysis of gel phase phosphatides, the steady-state initial velocities were hyperbolic functions of bulk lipid concentrations. At the 'pre-transition' temperature (34 degrees C for DPPC, 15 degrees C for DMPC), there was a large increase in the Michaelis parameter Vmax but no change in the parameter Km. A model was devised to account for these observations, in which the enzyme desorbs from the lipid surface after hydrolysis. The desorption rate constant is postulated to increase above the pretransition temperature. For hydrolysis of liquid crystalline phosphatides, the reaction consisted of a short initial burst of hydrolysis, a long 'lag' period of very slow reaction, followed by a dramatic increase in the reaction rate. Addition of 10 mol% lysolecithin or fatty acid abolished the 'lag' period. It was postulated that the enzyme adsorbs irreversibly to the surface of the liquid crystalline phase. Reaction products are postulated to stimulate desorption of enzyme from the surface. Thus, temperature-dependent changes in the rate of hydrolysis of dispersed phosphatidylcholines are attributed to changes in the rate of desorption of the enzyme from the lipid surface.
D055598:Chemical Phenomena; D002627:Chemistry, Physical; D003435:Crotalid Venoms / Q000378:metabolism*; D006868:Hydrolysis; D007700:Kinetics; D010713:Phosphatidylcholines / Q000378:metabolism*; D010740:Phospholipases / Q000378:metabolism*; D011663:Pulmonary Surfactants / Q000378:metabolism; D012995:Solubility; D013329:Structure-Activity Relationship; D013696:Temperature
D003435:Crotalid Venoms; D010713:Phosphatidylcholines; D011663:Pulmonary Surfactants; D010740:Phospholipases