Restoration of specific myxovirus receptors to asialoerythrocytes by incorporation of sialic acid with pure sialyltransferases.

Abstract

Removal of sialic acids from human erythrocytes with Vibrio cholera neuraminidase abolished hemagglutination by two paramyxoviruses, Sendai and Newcastle disease, and two orthomyxoviruses, a human influenza A2 virus (Japan/305/57), and an equine influenza A virus (Equine i/Prague/56). Selective replacement of sialic acid in the sequences NeuAccu2 + 6 Gal, NeuAca2 -+ GGalNAc, or NeuAccv2 + 3Gal was accomplished with three different homogeneous sialyltransferases, each of which is specific for the synthesis of one of these structures. Hemagglutination titers were restored to the original levels for all four viruses by the incorporation of about 40% of the sialic acid content of the native cells with the fl-galactoside a2 + 3 sialyltransferase. This enzyme has a strict acceptor substrate specificity for the Galfll + 3GalNAcal + 0-Thr/Ser sequence found in asialoglycophorin, the major erythrocyte glycoprotein, and forms the structure NeuAccvt -+ 3Gal/31+ 3GalNAcal -+ 0-Thr/Ser. Analysis of the labeled erythrocyte proteins by sodium dodecyl sulfategel electrophoresis showed that about 85 to 90% of the [l’C]NeuAc incorporated into erythrocytes co-migrated with glycophorin. Incorporation of sialic acid by other sialyltransferases did not result in hemagglutination by the Sendai, Newcastle disease, or equine influenza A viruses. In contrast, both of the other sialyltransferases restored hemagglutination by the human influenza A2 virus. Thus, attachment of sialic acid to erythrocytes in the sequence NeuAca2 -+ 6Gal or NeuAccvP + 6GalNAc restores erythrocyte receptor sites to only one of the four myxoviruses examined.