MOLECULAR-BASIS OF ALTERED RED-BLOOD-CELL MEMBRANE-PROPERTIES IN SOUTHEAST-ASIAN OVALOCYTOSIS - ROLE OF THE MUTANT BAND-3 PROTEIN IN BAND-3OLIGOMERIZATION AND RETENTION BY THE MEMBRANE SKELETON

Southeast Asian ovalocytosis (SAG) is an asymptomatic trait characteri zed by rigid, poorly deformable red cells that resist invasion by seve ral strains of malaria parasites. The underlying molecular genetic def ect involves simple heterozygous state for a mutant band 3 protein, wh ich contains a deletion of amino acids 400 through 408, linked with a Lys 56-to-Glu substitution (band 3-Memphis polymorphism). To elucidate the contribution of the mutant SAO band 3 protein to increased SAO re d blood cell (RBC) rigidity, we examined the participation of the muta nt SAO band 3 protein in increased band 3 attachment to the skeleton a nd band 3 oligomerization. We found first that SAO RBC skeletons retai ned more band 3 than normal cells and that this increased retention pr eferentially involved the mutant SAO band 3 protein, Second, SAO RBCs contained a higher percentage of band 3 oligomer-ankyrin complexes tha n normal cells, and these oligomers were preferentially enriched by th e mutant SAO protein. At the ultrastructural level, the increased olig omer formation of SAO RBCs was reflected by stacking of band 3-contain ing intramembrane particles (IMP) into longitudinal strands. The IMP s tacking was not reversed by treating SAO RBCs in alkaline pH (pH 11), which is known to weaken ankyrin-band 3 interactions, or by removing t he cytoplasmic domain of band 3 from SAO membranes with trypsin. Final ly, we found that band 3 protein in intact SAO RBCs exhibited a marked ly decreased rotational mobility, presumably reflecting the increased oligomerization and the membrane skeletal association of the SAO band 3 protein, We propose that the mutant SAO band 3 has an increased prop ensity to form oligomers, which appear as longitudinal strands of IMP and exhibit increased association with membrane skeleton. This band 3 oligomerization underlies the increase in membrane rigidity by preclud ing membrane skeletal extension, which is necessary for membrane defor mation. (C) 1995 by The American Society of Hematology.