BINDING OF NATURALLY-OCCURRING ANTIBODIES TO OXIDATIVELY AND NONOXIDATIVELY MODIFIED ERYTHROCYTE BAND-3

Both oxidative clustering (elicited by diamide treatment) and nonoxida tive clustering (elicited by zinc/BS3 (bis[sulfosuccinimidyl]suberate) treatment) of erythrocyte integral membrane proteins induce binding o f autologous antibodies with anti-band 3 specificity, followed by comp lement deposition and phagocytosis. Autologous antibodies eluted from nonoxidatively clustered erythrocytes bind to and stimulate phagocytos is of oxidatively damaged erythrocytes. Those eluted antibodies bind s pecifically to disulfide-crosslinked band 3 dimers generated by diamid e treatment. Band 3 dimerization and antibody binding are abrogated by cleavage of band 3 cytoplasmic domain. Thus, disulfide-crosslinked ba nd 3 dimers are the minimal band 3 aggregate with enhanced affinity fo r anti-band 3 antibodies. The eluted antibodies do not bind to band 3 dimers generated nonoxidatively by BS3 treatment but bind avidly to la rger band 3 clusters generated nonoxidatively by zinc/BS3 treatment. P ossibly, disulfide crosslinking of cytoplasmic domain cysteines induce s reorientation of intramembrane domains as to expose putative anti-ba nd 3 epitopes and allow bivalent binding of anti-band 3 antibodies. Ex tensive nonoxidative band 3 clustering appears to disrupt the native b and 3 conformation and generate reoriented dimers which expose putativ e anti-band 3 epitopes in the proper distance and orientation as to al low bivalent antibody binding.