Structural studies on the O-linked carbohydrate chains of human platelet glycocalicin.
Glycocalicin (140 kDa), constituting the main part of glycoprotein Ib (160 kDa), was released from the human platelet membrane by the action of a Ca2+-dependent protease, present in the platelet cytoplasm and liberated during sonication of the platelet suspension. After activation of the protease by Ca2+, the sonicated platelet suspension was subjected to differential centrifugation. The supernatant was applied to a column of wheat germ agglutinin linked to Sepharose 4B; glycocalicin was eluted from the column with 2.5% (w/v) N-acetylglucosamine. Glycocalicin was found to contain 40% carbohydrate by weight, representing N- as well as O-glycosidically linked carbohydrate chains. The O-glycosidic chains were split off by alkaline cleavage in the presence of 3H-labelled NaBH4. The liberated 3H-labelled oligosaccharide-alditols were fractionated on a DEAE-Sephadex A-25 column. The structures of the oligosaccharide-alditols were investigated by 500-MHz 1H-NMR spectroscopy. The major compound was identified as NeuAc alpha(2----3)Ga1 beta(1----3)[NeuAc alpha(2----3)Ga1 beta(1----4)GlcNAc beta(1----6)]GalNAc-ol. Two minor compounds were found to be NeuAc alpha(2----3)Gal beta(1----3)[NeuAc alpha(2----6)]GalNAc-ol and NeuAc alpha(2----3)Gal beta(1----3)GalNAc-ol.
D001792:Blood Platelets / Q000032:analysis*; D002241:Carbohydrates / Q000032:analysis*; D055598:Chemical Phenomena; D002621:Chemistry; D006023:Glycoproteins / Q000097:blood*; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008565:Membrane Proteins / Q000097:blood*; D019038:Platelet Glycoprotein GPIb-IX Complex; D011522:Protons
D002241:Carbohydrates; D006023:Glycoproteins; D008565:Membrane Proteins; D019038:Platelet Glycoprotein GPIb-IX Complex; D011522:Protons; C023592:glycocalicin