抄録
Binding specificity of naturally occurring anti-band 3 IgG antibody isolated from human plasma was investigated in a cell-free binding system. 125I-labeled human anti-band 3 IgG specifically bound to band 3 glycoprotein and lactoferrin, a glycoprotein that has poly-N-acetyllactosamine-type sugar chains like band 3, on the polyvinylidene difluoride blotting membrane. Binding was decreased by 50-70% when band 3 and lactoferrin were pretreated with N-glycosidase F, endo-β-galactosidase, or neuraminidase. Binding of 125I-anti-band 3 IgG to band 3-Sepharose gel was partially inhibited by band 3 oligosaccharides or lactoferrin, but was less inhibited by them after they had been treated with N-glycosidase F or endo-β-galactosidase. A significant part of 125I-anti-band 3 IgG that bound to the band 3-Sepharose gel was released upon treatment of the gel with N-glycosidase F or endo-β-galactosidase. IgG that binds to lactoferrin (anti-lactoferrin IgG) was isolated from normal human plasma. 125I-Anti-lactoferrin IgG bound to the band 3-Sepharose gel as effectively as to the lactoferrin-Sepharose. The antibody specifically bound to the band 3-and lactoferrin-blotted membrane depending on the poly-N-acetyllactosaminyl sugar chains of the blotted glycoproteins. The results indicate that a major part (about 70%) of anti-band 3 IgG recognizes the sialylated poly-N-acetyllactosaminyl sugar chains of band 3 and lactoferrin, and the remaining part (about 30%) of the antibody may recognize the polypeptide portion of band 3. This was supported by the observation that anti-band 3 IgG effectively bound to lactoferrin-Sepharose but 33% of the antibody did not. Anti-band 3 IgG with the carbohydrate-binding property was equally obtained whether fully denatured or barely denatured band 3 was used for isolation of anti-band 3 IgG by affinity chromatography. These results provide further evidence for our proposal that the binnding sites of anti-band 3 IgG to oxidized and senescent erythrocytes reside on the locally condensed sialylated poly-N-acetyllactosaminyl sugar chains of band 3 on the cell surface.
