Hydrophobic labeling of a single leaflet of the human erythrocyte membrane

The photoactivatable phospholipid 1-palmitoyl-2-[10-[4-[(trifluoromethyl)diazirinyl]phenyl]-[9-3H] -8-oxadecanoyl]-sn-glycero-3-phosphocholine [( 3H]PTPC) was synthesized with high specific radioactivity. When a sonicated dispersion of [3H]PTPC was incubated with human erythrocyte membranes (ghosts), the radiolabel was inserted spontaneously into the erythrocyte membrane. Photo-cross-linking of [3H]PTPC to membrane components and subsequent analysis of the distribution of radiolabel among polypeptide fragments of glycophorin allowed conclusions concerning the transbilayer distribution of [3H]PTPC in the erythrocyte membrane. Thus, [3H]PTPC was inserted exclusively into the outer leaflet of resealed ghosts, whereas with unsealed (leaky) ghosts, the photosensitive lipid was incorporated into both halves of the membrane simultaneously. These results are incompatible with fusion of [3H]PTPC liposomes with the erythrocyte membrane being responsible for the lipid transfer observed and suggest instead that PTPC exchanges spontaneously between membranes. This property of PTPC could be related to the critical micelle concentration of this lipid [(6 +/- 2) X 10(-9) M at 23 degrees C].