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Isolation and structural characterization of human lymphocyte and neutrophil gangliosides
Abstract:
Gangliosides were isolated from purified preparations of human peripheral blood lymphocytes and neutrophils. Structural analyses and comparisons were performed by direct probe mass spectrometry and by degradation studies with the following enzymes: Escherichia freundii endo-beta-galactosidase; Clostridium perfringens and Arthrobacter ureafaciens neuraminidase; and jack bean beta-N-acetylhexosaminidase and beta-galactosidase. This combination of techniques allowed us to obtain carbohydrate composition and sequence information without the aid of methylation or carbohydrate compositional analyses using only 1-2 mg of purified gangliosides. On the basis of these studies we propose that human lymphocytes and neutrophils have gangliosides with the following structures. NeuAc alpha 2 leads to 3Gal beta 1 leads to 4Glc beta 1 leads to 1Cer Structure A NeuAc alpha 2 leads to ? GlcNAc beta 1 leads to 3Gal beta 1 leads to 4Glc beta 1 leads to 1Cer Structure B NeuAc alpha 2 leads to ? Gal beta 1 leads to 3,4GlcNAc beta 1 leads to 3Gal beta 1 leads to 4Glc beta 1 leads to 1Cer Structure C All three compounds were isolated from both cell types with structure A being the major lymphocyte ganglioside and structure C the major neutrophil ganglioside. Structure B is a novel ganglioside and may represent a leukocyte-specific glycosphingolipid. Neuraminidase degradation studies demonstrated that only one ganglioside species of each cell type contains an internally linked sialic acid residue, and on the basis of thin layer chromatographic analysis this component is the same as the major brain ganglioside, GM1 (II3-N-acetylneuraminosyl-gangliotetraosylceramide). In addition, large gangliosides with the general structure NeuAc alpha 2 leads to ?(Gal beta 1 leads to 3,4GlcNAc beta 1 leads to 3)n Gal beta 1 leads to 4Glc beta 1 leads to 1Cer were isolated. These results are discussed as they relate to blood group antigens and specific cell surface markers in human leukocytes.