Oligosaccharide motion in erythrocyte membranes investigated by picosecond fluorescence polarization and microsecond dichroism of an optical probe

Abstract

Oligosaccharide chains on the surface of human erythrocytes were labeled with the probe eosin 5-thiosemicarbazide. The probe was conjugated to aldehydes produced by oxidation of sialic acid and galactose residues. The probe is associated mostly with glycophorin A after sialic acid labeling, whereas multiple components, including band 3 and lipids, are labeled after galactose oxidation. Fast molecular motion was studied by measuring steady-state and picosecond time-resolved fluorescence depolarization. Slower motions were investigated by observing flash-induced transient dichroism. It was found that both eosin-labeled sialic acid and galactose residues exhibit a rapid motion with correlation time of approximately 3 nsec. This motion is assigned to independent motion of the probe, r219w7 =motion, possibly in conjuction with a short segment of the oligosaccharide chain. The order parameter of the fast motion is 0.8-0.9, demonstrating that its angular amplitude is highly restricted. For eosin-labeled sialic acid, the order parameter in the microsecond time range is 0.2-0.3. It is deduced that a second, slower rotational motion is present, which is assigned to a cooperative motion of the oligosaccharide chains. The correlation time of this motion is in the range 10-7-10-5 sec. Some eosin-labeled galactose residues may have a similar slow mition, but most appear to be remarkably immobile over the time range 10-8-10-3 sec.