Biosynthesis and turnover of O-acetyl and N-acetyl groups in the gangliosides of human melanoma cells

Journal Article · Sun Aug 05 04:00:00 EDT 1990 · Journal of Biological Chemistry; (USA)

OSTI ID:6622389

Manzi, A E; Sjoberg, E R; Diaz, S; Varki, A

We and others previously described the melanoma-associated oncofetal glycosphingolipid antigen 9-O-acetyl-GD3, a disialoganglioside O-acetylated at the 9-position of the outer sialic acid residue. We have now developed methods to examine the biosynthesis and turnover of disialogangliosides in cultured melanoma cells and in Golgi-enriched vesicles from these cells. O-Acetylation was selectively expressed on di- and trisialogangliosides, but not on monosialogangliosides, nor on glycoprotein-bound sialic acids. Double-labeling of cells with (3H)acetate and (14C)glucosamine introduced easily detectable labels into each of the components of the ganglioside molecules. Pulse-chase studies of such doubly labeled molecules indicated that the O-acetyl groups turn over faster than the parent molecule. When Golgi-enriched vesicles from these cells were incubated with (acetyl-3H)acetyl-coenzyme A, the major labeled products were disialogangliosides. (Acetyl-3H)O-acetyl groups were found at both the 7- and the 9-positions, indicating that both 7-O-acetyl GD3 and 9-O-acetyl GD3 were synthesized by the action of O-acetyltransferase(s) on endogenous GD3. Analysis of the metabolically labeled molecules confirmed the existence of both 7- and 9-O-acetylated GD3 in the intact cells. Surprisingly, the major 3H-labeled product of the in vitro labeling reaction was not O-acetyl-GD3, but GD3, with the label exclusively in the sialic acid residues. Fragmentation of the labeled sialic acids by enzymatic and chemical methods showed that the 3H-label was exclusively in (3H)N-acetyl groups. Analyses of the double-labeled sialic acids from intact cells also showed that the 3H-label from (3H)acetate was exclusively in the form of (3H)N-acetyl groups, whereas the 14C-label was at the 4-position.

• OSTI ID: 6622389
• Journal Information: Journal of Biological Chemistry; (USA), Journal Name: Journal of Biological Chemistry; (USA) Vol. 265:22; ISSN JBCHA; ISSN 0021-9258
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
550901 -- Pathology-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ACETATES
ACETYLATION
ACYLATION
AMINES
ANIMAL CELLS
ANIMALS
BIOSYNTHESIS
CARBOHYDRATES
CARBON 14 COMPOUNDS
CARBOXYLIC ACID SALTS
CHEMICAL REACTIONS
CHROMATOGRAPHY
COENZYMES
DISEASES
DOUBLE LABELLING
GANGLIOSIDES
GLUCOSAMINE
GLYCOLIPIDS
HEXOSAMINES
HEXOSES
HYDROGEN COMPOUNDS
ISOTOPE APPLICATIONS
ISOTOPE DILUTION
LABELLED COMPOUNDS
LABELLING
LIPIDS
LIQUID COLUMN CHROMATOGRAPHY
MAMMALS
MAN
MELANOMAS
METABOLISM
MONOSACCHARIDES
NEOPLASMS
ORGANIC COMPOUNDS
PRIMATES
SACCHARIDES
SEPARATION PROCESSES
SIALIC ACID
SYNTHESIS
TRACER TECHNIQUES
TRITIUM COMPOUNDS
TUMOR CELLS
VERTEBRATES