Apolipoprotein-induced conversion of phosphatidylcholine bilayer vesicles into nanodisks.

摘要

Apolipoprotein mediated formation of nanodisks was studied in detail using apolipophorin III (apoLp-III), thereby providing insight in apolipoprotein-lipid binding interactions. The spontaneous solubilization of 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) vesicles occured only in a very narrow temperature range at the gel-liquid-crystalline phase transition temperature, exhibiting a net exothermic interaction based on isothermal titration calorimetry analysis. The resulting nanodisks were protected from proteolysis by trypsin, endoproteinase Glu-C, chymotrypsin and elastase. DMPC solubilization and the simultaneous formation of nanodisks were promoted by increasing the vesicle diameter, protein to lipid ratio and concentration. Inclusion of cholesterol in DMPC dramatically enhanced the rate of nanodisk formation, presumably by stabilization of lattice defects which form the main insertion sites for apolipoprotein alpha-helices. The presence of fully saturated acyl chains with a length of 13 or 14 carbons in phosphatidylcholine allowed the spontaneous vesicle solubilization upon apolipoprotein addition. Nanodisks with C13:0-phosphatidylcholine were significantly smaller with a diameter of 11.7 +/- 3.1nm compared to 18.5 +/- 5.6 nm for DMPC nanodisks determined by transmission electron microscopy. Nanodisk formation was not observed when the phosphatidylcholine vesicles contained acyl chains of 15 or 16 carbons. However, using very high concentrations of lipid and protein (>10mg/ml), 1,2,-dipalmitoyl-sn-glycero-3-phosphocholine nanodisks could be produced spontaneously although the efficiency remained low.