Phospholipase A(2) activity towards vesicles of DPPC and DMPC-DSPC containing small amounts of SMPC.
Phospholipase A(2) (PLA(2)) is an interfacially active enzyme whose hydrolytic activity is known to be enhanced in one-component phospholipid bilayer substrates exhibiting dynamic micro-heterogeneity. In this study the activity of PLA(2) towards large unilamellar vesicles composed of DPPC:SMPC and DMPC:DSPC:SMPC is investigated using fluorescence and HPLC techniques. Phase diagrams of the mixtures are established by differential scanning calorimetry and the PLA(2) activity, monitored by the lag time, is correlated with the phase behavior of the mixtures. In addition, the degree of lipid hydrolysis in the DMPC:DSPC:SMPC lipid mixtures is detected by HPLC. The PLA(2) activity is found to be significantly increased in the temperature range of the coexistence region where the lipid mixtures exhibit lateral gel-fluid phase separation. Furthermore, in the entire temperature range it is demonstrated that PLA(2) preferentially hydrolyzes the short chain DMPC lipid. This discriminative effect becomes less pronounced when the asymmetric lipid SMPC is present in the lipid substrate. Inclusion of SMPC into either DPPC or DMPC:DSPC vesicles prolongs the lag time. The results clearly show that the PLA(2) activity is significantly enhanced by lipid bilayer micro-heterogeneity in both one-component and multi-component lipid bilayer substrates. The PLA(2) activity measurements are discussed in terms of dynamic gel-fluid lipid domain formation due to density fluctuations and static lipid domain formation due to gel-fluid phase separation.
Høyrup
P
P
Department of Chemistry, Technical University of Denmark.
Mouritsen
O G
OG
Jørgensen
K
K
D015060:1,2-Dipalmitoylphosphatidylcholine / Q000737:chemistry*; D002152:Calorimetry, Differential Scanning; D002851:Chromatography, High Pressure Liquid; D004134:Dimyristoylphosphatidylcholine / Q000737:chemistry*; D008051:Lipid Bilayers / Q000737:chemistry*; D010316:Particle Size; D010713:Phosphatidylcholines / Q000737:chemistry; D010741:Phospholipases A / Q000737:chemistry*; D013050:Spectrometry, Fluorescence; D013696:Temperature; D013816:Thermodynamics; D013997:Time Factors
D008051:Lipid Bilayers; D010713:Phosphatidylcholines; D015060:1,2-Dipalmitoylphosphatidylcholine; C010942:1,2-distearoyllecithin; D010741:Phospholipases A; D004134:Dimyristoylphosphatidylcholine