Animal Glycoproteins Research Articles

Marker of peripheral blood granulocytes and monocytes of man recognized by two monoclonal antibodies VEP8 and VEP9 involves the trisaccharide 3‐fucosyl‐N‐acetyllactosamine

Two hybridoma antibodies (VEP8 and VEP9) raised against the promyelomonocytic leukemia cell line HL60 have previously been shown to distinguish human granulocytes and monocytes from other cells of the peripheral blood. We report here that both antibodies recognize the carbohydrate structure 3-fucosyl-N-acetyllactosamine with the following sequence: (formula; see text) This structure is the same as that recognized by a hybridoma antibody against mouse teratocarcinoma cells (anti-SSEA-1) which recognizes an early embryonic antigen in the mouse. Until recently this carbohydrate structure was considered to be rare among glycoproteins and glycosphingolipids. However, there is a growing list of human and animal glycoproteins in which this sequence has been detected by chemical and immunochemical methods. In this article we survey this information and discuss how this and other carbohydrate structures behave as differentiation- or tumor-associated antigens.

Chemistry and Biochemistry of d- and l-Fucose

This chapter discusses the various aspects of the chemistry of fucoses, especially the developments made in this field, and explains their metabolism and biochemistry while pointing out the possible biological functions and medical applications of fucoses and their compounds. Fucose is a 6-deoxyhexose having the “galacto” configuration—that is, it has cis-hydroxyl groups on C-3 and C-4. D-Fucose is usually found in plants and some microorganisms, in relatively simple glycosides comprising only a few sugar units. Some plant glycosides and polysaccharides crossreact with reagents for blood-group antigens, and interesting crossreactions has also been shown by methylated-fucose haptens and L-fucose-specific lectins. Great interest has been shown in the characterization of glycosyltransferases in connection with the studies on the tissue localization and mechanism of the biosynthesis of other animal glycoproteins. L-Fucose is found to inhibit the growth of some cells in culture, but only at high doses, and to inhibit the growth of implanted, mammary tumors in rats. The conversion of GDP-D-Man into GDP-Fuc serves as a preliminary stage in the biosynthesis of L-fucans, and its hydrolysis affords free L-fucose.

Isolation, physicochemical properties, and macromolecular composition of zona pellucida from porcine oocytes.

Oocytes released en masse from pig ovaries were isolated in large quantities by using sieving techniques. The isolated oocytes were gently homogenized, and the largely intact zona pellucida "ghosts" were purified by using sieving techniques. Sufficient amounts of zonae were recovered to permit, for the first time, adequate characterization of the zona pellucida in chemical, physical, and macromolecular terms. The isolated zonae were greater than 93% pure as determined by chemical, enzymatic, and microscopic criteria. The zonae were completely solubilized by a variety of conditions that do not break covalent bonds. The extent of solubilization was a function of pH, ionic strength, temperature, and the presence of various solubilizing agents such as detergents and urea. Chemically, the zonae were composed predominantly of protein (71%) and carbohydrate (19%). After acid hydrolysis of the zonae, no unusual amounts or types of amino acids were detected. The monosaccharides present after hydrolysis were those typically found in animal glycoproteins (Fuc, Man, Gal, GalNAc, and GlcNAc). Sialic acid in glycosidic linkage and sulfate and phosphate esters were present and were considered to be true constituents of the zona pellucida. Other substances detected, but considered contaminants rather than true constituents, included fatty acids (esterified and free) and uronic acids. The binding by several fluorescein-conjugated plant lectins to the in situ zona pellucida was determined by using light microscopy. The binding of the lectins to the zona pellucida was not uniform, indicating that the carbohydrate moieties of the zona pellucida were asymmetrically distributed. The zona pellucida was composed of at least three macromolecules as indicated by immunodiffusion and sodium dodecyl sulfate gel electrophoresis experiments. Determination of the number of macromolecules composing the zona pellucida was compromised by the aggregation and/or microheterogeneity of its constituent macromolecules. We conclude that the zona pellucida is composed of several glycoprotein macromolecules; interaction of these macromolecules to form supramolecular complexes and the integral zona pellucida is dependent on noncovalent forces.

Analysis of methyl glycosides as their trimethylsilyl ethers: on-columns Re-N-acetylation and improved gas— liquid chromtographic separation

A rapid procedure is described for methyl glycoside derivatization for gas— liquid chromatographic analysis. Re-N-acetylation is carried out after the trimethylsilylation step directly in a gas chromatographic column by injecting acetic anhydride together with a sample. A quantitative acetylation of amino groups occurs, whereas trimethysilylated hydroxyls remain intact. The entire procedure of treating a glyco- protein hydrolysate prior to the analysis takes ca. 40 min. the use of a 3.5-m column with OV-1 as stationary phase provides a complete peak separation for the derivatives of all of the neutral and amino sugars and N-acetylneuraminic acid commonly encountered in animal glycoproteins, and the analysis can be performed within 2.5 h.

Human pathologic IgM glycopeptides. Evidence for a very high mannose content glycopeptide

The carbohydrate composition of a human IgM myeloma protein has been studied and a particularly unusual glycopeptide purified and partially characterized. It contains 10 mannose residues and 2 N-acetylglucosamine residues in its oligosaccharide unit, N-glycosidically linked to the peptide structure of the heavy chain. Such a high quantity of mannose is not often found in the carbohydrate chains of animal glycoproteins and suggests an unusual process during the immunoglobulin glucidic chain biosynthesis.

The formation of glycoproteins in tissues of higher plants. Specific labelling with D-(1- 14 C)glucosamine.

1. Radioactivity from d-[l-(14)C]glucosamine is incorporated into ethanol-insoluble compounds of high molecular weight in a number of plant tissues, including roots of corn (Zea mays), callus cells of sycamore (Acer pseudoplatanus), axenic cultures of duckweed (Lemna minor) and germinating seedlings of corn, broad bean (Vicia faba) and barley (Hordeum vulgare). 2. Except in the case of Lemna, where some of the radioactivity was recovered in glucose, hydrolysis of these ethanol-insoluble materials with acid released [(14)C]glucosamine as the major radioactive product. 3. The labelled compounds isolated from Zea roots and the Acer cells are believed to be glycoproteins rather than polysaccharides on the basis of their solubility properties, their charge characteristics and their susceptibility to hydrolysis by 0.5m-potassium hydroxide and by the proteases trypsin and Pronase. Further, radioactive peptides were isolated and purified after Pronase treatment and shown to contain glucosamine as well as a number of amino acids. 4. The experiments therefore indicate that d-[(14)C]glucosamine can be used as a specific precursor of the amino sugar units of plant as well as animal glycoproteins.