Purification of Forssman and human blood group A glycolipids by affinity chromatography on immobilized Helix pomatia lectin.
A method for the affinity purification of intact glycolipids having nonreducing terminal alpha 1-3 linked N-acetylgalatosamine residues has been developed. This technique relies on the retention of the carbohydrate-binding specificity of immobilized Helix pomatia lectin in aqueous solutions of tetrahydrofuran. Both Forssman glycolipid and a mouse blood group A-active hexaosylceramide were bound by columns of the lectin equilibrated in a solvent containing 95% tetrahydrofuran and 5% water. After application of a step gradient of increasing water content up to 50%, the specifically bound glycolipids were eluted in solvent containing N-acetylgalactosamine. The Forssman and A-active glycolipids were similarly purified in a single chromatographic step from total lipid extracts of sheep and human type A erythrocyte stroma, respectively. Nonspecifically bound lipids and glycolipids were eluted from this column by simply increasing the water content of the eluting buffer. The extension of this method to other carbohydrate-binding proteins including lectins and monoclonal antibodies may provide a rapid purification of glycolipids based on their carbohydrate structures.
Torres
B V
BV
Department of Biochemistry and Nutrition, Virginia Tech, Blacksburg 24061.
Smith
D F
DF
D000818:Animals; D001789:Blood Group Antigens / Q000032:analysis*; D002846:Chromatography, Affinity; D004912:Erythrocytes / Q000032:analysis; D005915:Globosides / Q000032:analysis*; D006017:Glycolipids / Q000097:blood*; D006028:Glycosphingolipids / Q000032:analysis*; D006801:Humans; D037102:Lectins; D008055:Lipids / Q000097:blood; D051379:Mice; D011485:Protein Binding; D012756:Sheep
D001789:Blood Group Antigens; D005915:Globosides; D006017:Glycolipids; D006028:Glycosphingolipids; C038095:Helix lectin; D037102:Lectins; D008055:Lipids; C010163:Forssman glycolipid