Phospholipid translocation in various bilayr membranes using radioactive phospholipid probes

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Four kinds of radioactive phospholipid probes (phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, phosphatidic acid) were synthesized. The probes, termed C5-PC,C5-PE,C5-PS,C5-PA,respectively) have 14C labelled pentanoyl group at the second position and palmitoyl or stearoyl group at the first position of the glycerol skeleton of the phospholipids. The probes were applied to the measurement of phospholipid translocation in human erythrocyte plasma membranes. The results were consistent with those obtained with spin-labelled phospholipid probes. The probes were revealed to be useful for the measurement of phospholipid translocation across lipid bilayr membranes with high sensitivity.Effect of X-irradiation on the aminophospholipid translocation in erythrocyte membranes was measured. The initial velocity of PS-translocation was increases a little by the irradiation of 50 Gy, however, the plateau value was not affected. In this experimental condition, hemolysis and lipid peroxidation were minimal.Three kinds of the probes (C5-PC,C5-PE,C5-PS) were used to measure the phospholipid translocation in the plasma membrane of synaptosomes prepared from electric organ of Narke japonica. Phosphatidylserine-specific and ATP-dependent translocation was observed. The property of the translocator is different from that of erythrocyte membrane with repect to phospholipid specificity.The phospholipid translocation in synaptic vesicle membranes prepared from electric organ of Narke japonica was also measured. In this experiment, I have used spin-column technique to separate quickly the synaptic vesicles and bovine serum albumin which extracts the probe from outer leaflet of the bilayr. There was a translocator which has no phospholipid specificity and no ATP requirement.