Effects of physical states of phospholipids on the incorporation and cytochalasin B binding activity of human erythrocyte membrane proteins in reconstituted vesicles

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Abstract
Proteoliposomes were reconstituted from a Triton extract of human erythrocyte membrane proteins and a mixture of phosphatidylcholine (PC) and phosphatidylethanolamine (PE) of varying ratios. With mixtures of egg PC and soybean PE, the protein/lipid ratio of the reconsituted vesicles was maiximal at 25% PC and 75% PE, the composition which is known to have a maximum bilayer disruption (higest occurrence of lipidic particles seen by freeze-fracture electron microscopy). With mixtures of 1-palmitoyl-2-oleoyl-PC and dilinoleoyl-PE, which give vesicles with few isolated lipid particles at room temperature, the effect was less pronounced. The specific activity of the cytochalasin B(CB) binding protein in the reconstituted vesicles, on the other hand, was increased monotonically up to severalfold as the PC content was increased in the egg PC/soybean PE mixture. A similar increase was observed when soybean PE was partially substituted by dimyristoyl-PC, cholesterol or transphosphatidylated PE from egg PC. Evidently, reexisting defects in the lipid bilayer promote protein incorporation into the bilayer during reconstitution whereas reduction of the bilayer fluidity facilitates the CB binding activity in the reconstituted vesicles.