Analytical detection of 9(4)-O-acetylated sialoglycoproteins and gangliosides using influenza C virus.
The unique glycoprotein of influenza C virus, designated hemagglutinin (HEF), exhibits three functions: hemagglutination, esterase activity, and fusion factor. As the virus uses 9-O-acetylated sialic acid as a high-affinity receptor determinant for attachment to cells, its binding activity was used to reveal O-acetylated sialic acid residues after polyacrylamide gel electrophoresis and transfer onto nitrocellulose sheets of proteins and thin-layer chromatography of lipids. The specificity of the binding for O-acetylated sialoglycoconjugates was investigated. Our results showed that influenza C virus could detect the different forms of the two murine glycophorins which are known to be O-acetylated sialoglycoconjugates. The virus also bound to O-acetylated gangliosides isolated from embryonic chicken brain such as purified O-acetylated NeuAc alpha (2-8)NeuAc alpha (2-8)NeuAc alpha (2-3)Gal beta (1-4)Glc beta (1-1)ceramide (GT3). The esterase activity of the HEF protein of influenza C virus was used to unmask the sialic acid. After its deacetylation by the virus enzyme, the O-acetylated GT3 was recognized by a monoclonal antibody which binds only to the nonacetylated derivative. The results presented here show that influenza C virus is a discriminating analytical probe for identifying O-acetylated sialoglycoconjugates directly after Western blotting of proteins and thin-layer chromatography of lipids, thus providing a new analytical tool.
Manuguerra
J C
JC
Unité d'écologie virale, laboratoire de la grippe, Institut Pasteur, Paris, France.
DuBois
C
C
Hannoun
C
C
D000107:Acetylation; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000956:Antigens, Viral / Q000276:immunology; D015153:Blotting, Western; D002642:Chick Embryo; D003101:Collodion; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes; D004910:Erythrocyte Membrane / Q000378:metabolism; D004950:Esterases / Q000378:metabolism; D005732:Gangliosides / Q000032:analysis* / Q000378:metabolism; D009982:Gammainfluenzavirus / Q000201:enzymology / Q000276:immunology / Q000378:metabolism*; D051379:Mice; D012756:Sheep; D012795:Sialoglycoproteins / Q000032:analysis* / Q000378:metabolism
D000911:Antibodies, Monoclonal; D000956:Antigens, Viral; D000939:Epitopes; D005732:Gangliosides; D012795:Sialoglycoproteins; D003101:Collodion; D004950:Esterases