Glycosphingolipid binding specificities of rotavirus: Identification of a sialic acid-binding epitope

The glycosphingolipid binding specificities of neuraminidase-sensitive (sim ian SA11 and bovine NCDV) and neuraminidase-insensitive (bovine UK) rotavir us strains were investigated using the thin-layer chromatogram binding assa y. Both triple-layered and double-layered viral particles of SA11, NCDV, an d UK bound to nonacid glycosphingolipids, including gangliotetraosylceramid e (GA1; also called asialo-GM1) and gangliotriaosylceramide (GA2; also call ed asialo-GM2). Binding to gangliosides was observed with triple-layered pa rticles but not with double-layered particles. The neuraminidase-sensitive and neuraminidase-insensitive rotavirus strains showed distinct ganglioside binding specificities. All three strains bound to sialylneolactotetraosylc eramide and GM2 and GD1a gangliosides, However, NeuAc-GM3 and the GM1 gangl ioside were recognized by rotavirus strain UK but not by strains SA11 and N CDV, Conversely, NeuCc-GM3 was bound by rotaviruses SA11 and NCDV but not b y rotavirus UK. Thus, neuraminidase-sensitive strains bind to external sial ic acid residues in gangliosides, while neuraminidase-insensitive strains r ecognize gangliosides with internal sialic acids, which are resistant to ne uraminidase treatment. By testing a panel of gangliosides with triple-layer ed particles of SA11 and NCDV, the terminal sequence sialyl-galactose (NeuC c/NeuAc alpha3-Gal beta) was identified as the minimal structural element r equired for the binding of these strains, The binding of triple-layered par ticles of SA11 and NCDV to NeuGc-GM3, but not to NeuAc-GM3, suggested that the sequence Neucc alpha 3Gal beta is preferred to NeuAc alpha 3Gal beta, F urther dissection of this binding epitope showed that the carboxyl group an d glycerol side chain of sialic acid played an important role in the bindin g of such triple-layered particles.