Reorientation rates and asymmetry of distribution of lysophospholipids between the inner and outer leaflet of the erythrocyte membrane.

Labelled lysophospholipids were inserted into the outer layer of the erythrocyte membrane and their reorientation (flip) to the inner layer quantified by following the increase of the fraction of lysophospholipids not extractable by albumin. Flip rate constants were calculated from the kinetics of equilibration of the lysophospholipids between two compartments, the outer and the inner leaf of the bilayer, in the early phase of the flip kinetics where correction for non-enzymatic hydrolysis and acylation could be omitted. The distribution of a lysophospholipid finally attained reflects its affinity for the two layers. Whereas lysophosphatidylcholine has a slight preference for the outer layer of the membrane, lysophosphatidylserine spontaneously concentrates in the inner layer up to a ratio of 4:1. This asymmetry mimics the distribution of phosphatidylserine in the native membrane. Flip rates depend on membrane lipid compositions. They are enhanced by cholesterol depletion. Comparison of various mammalian species demonstrates that erythrocytes with a higher phosphatidylcholine/sphingomyelin ratio and high content of polyunsaturated fatty acids (mouse and rat) have a high transbilayer mobility, in contrast to erythrocytes with a low phosphatidylcholine/sphingomyelin ratio and a low content of polyunsaturated fatty acids (ox). Molecular properties of lysophospholipids influence their transbilayer mobility. Flip rates of lysophospholipids are enhanced not only by unsaturation of their fatty acid, but also by a negative net charge on the headgroup. This indicates that the strongly asymmetric distribution of phosphatidylserine in the native erythrocyte membrane, which is maintained for the lifespan of the cell, does not result from a lack of transbilayer mobility.