Forssman Glycolipid Research Articles

Determination of position of substitution on 2-acetamido-2-deoxy- D-glucosyl residues in glycolipids

The substitution site on 2-acetamido-2-deoxy-D-galactosyl residues in oligosaccharide chains of glycolipids was determined by permethylation of the glycolipid with methyl iodide in the presence of dimethylsulfinyl carbanion, methanolysis of the permethylated product under mild conditions, acetylation with acetic anhydride-pyridine, and identification of the resulting substituted methyl glycosides of 2-deoxy-2(N-methylacetamido)-D-galactose by g.l.c. The method was applied to glycolipids of known structure, including normal brain ganglioside, Tay-Sachs ganglioside, and Forssman glycolipid.

Determination of position of substitution on 2-acetamido-2-deoxy- D-galactosyl residues in glycolipids

The substitution site on 2-acetamido-2-deoxy- D-galactosyl residues in oligosaccharide chains of glycolipids was determined by permethylation of the glycolipid with methyl iodide in the presence of dimethylsulfinyl carbanion, methanolysis of the permethylated product under mild conditions, acetylation with acetic anhydride-pyridine, and identification of the resulting substituted methyl glycosides of 2-deoxy-2-( N-methylacetamido)- D-galactose by g.l.c. The method was applied to glycolipids of known structure, including normal brain ganglioside, Tay—Sachs ganglioside, and Forssman glycolipid.

Characterization of the Forssman Glycolipid Hapten of Horse Kidney by Mass Spectrometry

Abstract The Forssman glycolipid hapten was isolated from horse kidney and analyzed by direct inlet mass spectrometry of undergraded lipid derivatives. Methylated glycolipid gave no molecular ions, but some important carbohydrate sequence and ceramide (fatty acid and long chain base) ions. After reduction of this derivative with LiAlH4 (amide group of ceramide and of amino sugars were converted to the corresponding amines) a molecular weight ion (at m/e 1782) was recorded, as well as ions which were conclusive for the type and exact ratio of sugars and their sequence. Without any conventional degradative studies it was possible to conclude that the glycolipid was a pentaglycosyl-ceramide with the sequence hexosamine-hexosamine-hexose-hexose-hexose-ceramide, as first proposed by B. Siddiqui and S. Hakomori ((1971) J. Biol. Chem. 246, 5766–5769). The fatty acids were mainly nervonic (24:1) and lignoceric (24:0) acid but also C16 to C23 fatty acids. Both sphingosine and phytosphingosine were present. Evidence was obtained from the spectra for a preferential combination of long chain (24) fatty acid and phytosphingosine.

Use of mass spectrometry for the carbohydrate composition and sequence analysis of glycosphingolipids.

Mass spectrometry has been applied for a detailed structural characterization of glycosphingolipids. Methylated and methylated plus reduced glycolipids (amide groups of ceramide and amino sugars were reduced to the corresponding amines) were found the most useful derivatives. Molecular weight ions [M – 1] were not obtained for methylated lipids with more than four sugar units, but were easily obtained for reduced derivatives, so far with six sugars. Spectra of reduced derivatives showed very abundant ions containing the complete carbohydrate and the fatty acid, allowing a conclusion to be made concerning the carbohydrate composition (ratio of hexose and hexosamine and sialic acid). In addition, conclusive information was obtained for the carbohydrate sequence, including branching of the chain. By a combined use of both derivatives the long chain base and fatty acid composition was also evident. Spectra are discussed for a synthetic monoglycosylceramide, the Forssman glycolipid hapten (pentaglycosylceramide) and two gangliosides (a triglycosylceramide and a pentaglycosylceramide). The mass spectrum of the Forssman glycolipid provides the first conclusive evidence for two amino sugars in the terminal disaccharide of the molecule.

Structure of a Pentahexosylceramide (Forssman Hapten) from Canine Intestine and Kidney

A pentahexosylceramide isolated from canine kidney and intestine was shown by enzyme hydrolyses, partial acid hydrolysis, permethylation and analysis of the products by gas-liquid chromatography-mass spectrometry, and serological tests to be a Forssman hapten identical in structure with the Forssman glycolipid from horse spleen. The assigned structure was N-acetylgalactosaminyl(α1→3) N-acetylgalactosaminyl(β1→3)galactosyl(α1→4)galactosyl(β1→4)glucosyl-(1→1)ceramide. In the course of this work, mass spectrometry was employed to elucidate the structures of partially methylated alditol acetates derived from hexosamines.

The effect of dibutyryl adenosine 3′:5′-cyclic monophosphate on the synthesis of glycolipids by normal and transformed Nil cells

The synthesis of complex glycolipids was examined in two clones (Nil 1Cl and Nil 2Cl) of the Nil 2 line and their hamster sarcoma virus-transformed derivatives. Nil 2Cl contains three neutral glycolipids which increase in concentration as cells become confluent. However, the kinetics of increase differs for each of the three. The Forssman glycolipid increases in parallel with cell growth and reaches a maximal level just before cells reach their saturation density. Synthesis of globoside increases significantly only after the culture reaches saturation density. These “density-dependent” glycolipids are lost or decreased in concentration after transformation by hamster sarcoma virus. The growth of both normal and transformed cells is inhibited by treatment with dibutyryl cyclic AMP. The density at which they stop growth is dependent upon the inoculum density. The glycolipid patterns of normal and transformed cells has been examined in the presence or absence of dibutyryl cyclic AMP. In the presence of the nucleotide transformed cells did not regain the glycolipids or density-dependent synthesis of glycolipids which they lost as a result of transformation.

Contact-dependent enhancement of net synthesis of forssman glycolipid antigen and hematoside in nil cells at the early stage of cell-to-cell contact

Contact-dependent enhancement of net synthesis of forssman glycolipid antigen and hematoside in nil cells at the early stage of cell-to-cell contact