Asymmetric lateral mobility of phospholipids in the human erythrocyte membrane.

Abstract

The fluorescent phospholipid 1-acyl-2-[12-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)aminododecanoyl]phosphatidylcholine (NBD-phosphatidylcholine) and the corresponding aminophospholipid derivatives (NBD-phosphatidylethanolamine and NBD-phosphatidylserine) were introduced in the human erythrocyte membrane by a nonspecific phospholipid exchange protein purified from corn. The lateral mobility of the fluorescent phospholipids was measured by using an extension of the classical photobleaching recovery technique that takes advantage of a modulated fringe pattern and provides a high sensitivity. In intact erythrocytes and in ghosts resealed in the presence of ATP, the fluorescence-contrast curves after photobleaching decayed biexponentially corresponding to two lateral diffusion constants. With NBD-phosphatidylcholine, the majority of the signal corresponded to a "slow" component (1.08 x 10-9 cm2/sec at 20°C), whereas with the amino derivatives the majority of the signal corresponded to a "fast" component (5.14 x 10-9 cm2/sec at 20°C). If the ghosts were resealed without ATP, the fast component of the aminophospholipids disappeared. We interpret these results as follows: (i) Provided the cells or the ghosts contain ATP, the three fluorescent phospholipids distribute spontaneously between inner and outer leaflets as endogenous phospholipids, namely NBD-phosphatidylcholine is located in the outer leaflet, while both aminophospholipids are preferentially located in the inner leaflet. (ii) The viscosity of the inner leaflet of human erythrocyte membranes is lower than that of the outer leaflet.ESR as well as fluorescence techniques have been used to assess the physical properties of the erythrocyte membrane. Several independent workers have suggested that the outer monolayer has a higher viscosity (1-3). However, the fluorescence polarization experiments by Schachter et al. (4) suggested a lower viscosity of the outer monolayer. Photobleaching experiments in turkey erythrocytes by Henis et al. (5) indicated identical lipid diffusibility in both layers below 24°C and higher lipid diffusibility in the external layer above 30°C. However, the same group reported a higher lipid diffusibility in the internal monolayer of human erythrocytes at 20°C and a higher lipid diffusibility in the external monolayer at 6°C (6). Thus the difference in lipid "mobility" or "fluidity" between inner and outer leaflets is still controversial.There are several difficulties with these experiments. First, the selective probing of the inner and outer monolayer could introduce spurious effects. Henis and collaborators (5, 6) attempted to compare nonpermeant probes in intact cells and in leaky ghosts. The drawback of their method was to compare an intact cell with a modified system and assume arbitrarily that the membrane fluidity was not affected. An alternative approach is to compare the results obtained with selective probes ofthe inner or outer monolayer. In the native erythrocyte membrane, phosphatidylcholine (PtdCho) is located in the outer mo...

Comparative study of the lateral motion of extrinsic probes and anthracene‐labelled constitutive phospholipids in the plasma membrane of Chinese hamster ovary cells

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Weak interaction of spectrin with phosphatidylcholine‐phosphatidylserine multilayers: A 2H and 31P NMR study

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Phosphatidylserine, a death knell

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