Transbilayer mobility of phospholipids in the erythrocyte membrane. Influence of the membrane skeleton

Oxidative treatment of erythrocytes results in a strong enhancement of transbilayer reorientation of exogenous lysophospholipids. Only upon selective oxidation of SH-groups and concomitant crosslinking of spectrin by diamide, however, does asymmetry of distribution of inner layer phospholipids, phosphatidylethanolamine and phosphatidylserine in the erythrocyte membrane become lost. This indicates that asymmetry is not due to a low transbilayer reorientation of the inner layer phospholipids, but that phosphatidylethanolamine and phosphatidylserine do not have access to the flip sites in the native membranes. A role of membrane skeleton proteins in the suppression of access of inner layer phospholipids to the flip sites is indicated by the loss of asymmetry of both phospholipids in spectrin depleted inside out vesicles and in vesicles produced by heat denaturation of spectrin. This idea is further supported by the observation that inner layer phospholipid analogues lysophosphatidylethanolamine and lysophosphatidylserine spontaneously accumulate in the inner layer of native erythrocytes, whereas lysolecithin does not. Moreover, this asymmetry is abolished in inside out vesicles.