Publisher Summary Lectins play an important role in the development of immunology. Lectins also find application in serological laboratories for typing blood and determining secretor status, separating leucocytes from erythrocytes, and agglutinating cells from blood in the preparation of plasma. They serve as reagents for the detection, isolation, and characterization of carbohydrate-containing macromolecules, including blood-group antigens. In their interaction with saccharides, lectins serve as models for carbohydrate-specific antibodies, with the important advantage to purify lectins in gram quantities. Lectins are classified according to their carbohydrate-binding specificity that includes D-mannose(D-glucose)-binding lectins and 2-acetamido-2-deoxy-D-glucose-binding lectins. The chapter considers only those lectins that have been purified to homogeneity, and studied with regard to their biophysical, biochemical, and carbohydrate-binding specificity. The chapter also describes the cell-binding and biological properties of lectins. The chapter concludes with the description of several glycopeptide structures showing the carbohydrate-binding loci with which various lectins interact.

The lectins : carbohydrate-binding proteins of plants and animals

Publisher Summary This chapter discusses the interactions of lectins with animal cell surfaces. Lectin interactions with cell surfaces can be put into a proper framework for understanding complex phenomena such as lectin-induced mitogenic stimulation, cell agglutination, and lectin-mediated cell toxicity. Lectins have been isolated from a wide variety of plants and animals, from legumes to horseshoe crabs. Lectins generally have the property of agglutinating cells through their cell surface oligosaccharide determinants, and in some cases these determinants have been elucidated by determining the sequences or sugars responsible for saccharide inhibition of agglutination. Because of the carbohydrate-binding specificities of lectins, some of these agglutinins have proven to be quite useful for clinical blood typing and structural studies of blood group substances, in analysis of the surface structure of normal and tumor cells, for specific isolation of glycoproteins and oligo- and mucopolysaccharides, in studies on mitogenesis, as antigen-antibody models, and so on. They have also proven to be invaluable as specific molecular probes for studying membrane, cell, and tissue structure and organization.

The interactions of lectins with animal cell surfaces.

The interaction of wheat germ agglutinin with sialoglycoproteins. The role of sialic acid.

A comparative study of glycoproteins from the surface of control and Rous sarcoma virus transformed hamster cells.

Isolation of the Receptors for Wheat Germ Agglutinin and the Ricinus communis Lectins from Human Erythrocytes Using Affinity Chromatography

Novikoff ascites tumor cells were found to be agglutinable by both concanavalin A and wheat germ agglutinin (WGA). A sialoglycopeptide-containing fraction, released by papain digestion of Novikoff tumor cell suspensions, inhibits the agglutination reactions, indicating that receptor sites for these plant lectins are components of this glycopeptide fraction. The purpose of this research was to develop methods for separation of the glycopeptide fraction into its component molecules and to characterize these components regarding their ability to inhibit agglutination of Novikoff tumor cells by concanavalin A and WGA.

The sialoglycopeptide-containing fraction was digested with Pronase to minimize variation in the peptide chain length of the component glycopeptides. The Pronase-digested glycopeptide fraction was submitted to gel filtration on Sephadex G-50 and subsequently to ion-exchange chromatography on DEAE-cellulose. Gel filtration resolved the glycopeptides into two molecular weight classes, a low-molecular-weight fraction (M.W. 2000 to 3300) and a higher-molecular-weight fraction (M.W. >3300). This latter fraction consists of molecules of widely different molecular weights, some of the material being excluded from Sephadex G-100. Ion-exchange chromatography of the low-molecular-weight material resolved four sialoglycopeptide fractions which inhibited agglutination of Novikoff tumor cells by concanavalin A but which were inactive in inhibiting agglutination by WGA. Ion-exchange chromatography of the higher-molecular-weight material resulted in the isolation of a sialoglycopeptide fraction which possessed high specificity as an inhibitor of agglutination by WGA.

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Isolation of tumor cell surface binding sites for concanavalin A and wheat germ agglutinin.

Summary Sialic acid has been shown to contribute significantly to the cell-surface charge of tumor cells. In order to elucidate the chemical nature of the sialic acid-containing molecules present on the tumor cell surface, a procedure for the isolation of a sialoglycopeptide fraction from the surface of Novikoff ascites cells has been developed. Digestion of tumor cell suspensions with papain liberates a sialoglycopeptide fraction from the cell surface. This fraction was partially purified using trichloroacetic acid precipitation, dialysis, and gel filtration on Sephadex G-50. The sialoglycopeptide fraction contains 65–80% of the neuraminidase-labile sialic acid present on the surface of these cells. Quantitative analysis of the composition of this fraction revealed the presence of the following monosaccharides, expressed per mg glycopeptide: 0.39 µmole sialic acid, 0.41 µmole glucosamine, 0.23 µmole galactosamine, 0.15 µmole mannose, 0.54 µmole galactose, 0.53 µmole glucose, and 0.15 µmole uronic acid. Amino acid analysis indicated a peptide content of 15%. Amino acids present in the highest amounts were threonine, aspartic acid, glutamic acid, serine, proline, alanine, and valine (molar ratio, 4:2:1.5:1:1:1:1).

https://cancerres.aacrjournals.org/content/canres/29/11/2034.full.pdf

Isolation and Chemical Characterization of a Cell-Surface Sialoglycopeptide Fraction from Novikoff Ascites Cells

The biochemical composition of any isolated organelle is affected by the agent(s) to which it has been exposed during the isolation and purification procedures. Many agents are known to extract molecular components either selectively or nonselectively. The protein composition of isolated chromosomes exposed to fluorochromes must be examined before proceeding to the biochemical analysis of the proteins from individual chromosomes purified by flow cytometry techniques. We have examined the polyacrylamide gel electrophoresis protein profiles from from metaphase chromosome exposed to the fluorochromes and staining conditions commonly used for flow cytometry. We find that the electropherograms of the protein of metaphase chromosomes which have been exposed to various fluorochromes are indistinguishable from controls. The analysis of proteins from individual chromosomes sorted by flow cytometry can now be undertaken.

Virginia P. Wray

Papers

Isolation of tumor cell surface binding sites for concanavalin A and wheat germ agglutinin.

Isolation and Chemical Characterization of a Cell-Surface Sialoglycopeptide Fraction from Novikoff Ascites Cells

Proteins from metaphase chromosomes treated with fluorochromes.

Selective solubilization and purification of cell-surface concanavalin A-binding glycoproteins from Novikoff hepatocellular carcinoma cells