Association of GM4 Ganglioside with the Membrane Surrounding Lipid Droplets in Shark Liver

Abstract

By TLC, GM4 was found to be the major ganglioside in the liver of six shark species examined: Odontaspis taurus, Negaprion brevirostris, Sphyrna lewini, Mustelus griseus, Mustelus manazo, and Prionace glauca. A detailed analysis of the glycosphingolipids (GSLs) in the liver of O. taurus (sand tiger shark) showed that it contained approximately 110 nmol of lipid-bound sialic acid per gram of wet tissue, of which 80% was GM4. By extracting the liver of O. taurus with chloroform/methanol, followed by chromatographic separation of GSLs using DEAE-Sephadex A-25 and Iatrobeads columns, we have isolated GM4 in pure form with a yield of approximately 5 mg per 100 g of wet tissue. The structures of both the sugar chain and the ceramide moiety of this GM4 were analyzed by chemical analysis, mass spectrometry, and NMR spectroscopy. Similar to GM4 isolated from other sources, 92% of fatty acids in the ceramide of this GM4 were 2-hydroxylated. However, unlike the long-chain bases found in other GSLs, the total long-chain bases in this GM4 were found to contain 43% octadecasphingenine and 50% nonadecasphingenine.Immunohistochemical analysis using a monoclonal antibody against GM4 revealed that the hepatocytes of both M. griseus (spotless smooth hound) and M. manazo (smooth hound) were filled with lipid droplets and GM4 was primarily associated with the membrane structure surrounding lipid droplets.

Keywords

• NMR spectroscopy
• secondary ion mass spectrometry
• collision-induced dissociation mass spectrometry