Characterization of a xylose-specific antiserum that reacts with the complex asparagine-linked glycans of extracellular and vacuolar glycoproteins

• Published in: Plant physiology (Bethesda) 1989-07, Vol.90 (3), p.1182-1188
• Main Authors: Michel Laurière, Christiane Laurière, Chrispeels, Maarten J., Johnson, Kenneth D., Sturm, Arnd
• Format: Article
• Language: English
• Subjects: Analysis of the immune response. Humoral and cellular immunity; Antibodies; anticorps; anticuerpos; Antiserum; Biological and medical sciences; Cell walls; Cellular and Structural Biology; cotiledones; cotyledon; cotyledons; daucus carota; enzimas; enzyme; enzymes; Epitopes; Fundamental and applied biological sciences. Psychology; Fundamental immunology; Genetics; glicoproteinas; Glycopeptides; glycoproteine; Glycoproteins; graine; immune serum; Immunobiology; immunserum; lectina; lectine; lectins; Life Sciences; nicotiana tabacum; Oligosaccharides; Organs and cells involved in the immune response; pared celular; paroi cellulaire; phaseolus vulgaris; Phytohemagglutinins; Plants; Plants genetics; Polysaccharides; proteinas; proteine; proteins; saccharomyces cerevisiae; seeds; semilla; suero inmune
• ISSN: 0032-0889; 1532-2548
• DOI: 10.1104/pp.90.3.1182

Antibodies were raised against carrot (Daucus carota) cell wall beta fructose that was either in a native configuration (this serum is called anti-beta F1) or chemically deglycosylated (anti-beta F2). The two antisera had completely different specificities when tested by immunoblotting. The anti-beta F1 serum reacted with beta fructosidase and many other carrot cell wall proteins as well as with many proteins in extracts of bean (Phaseolus vulgaris) cotyledons and tobacco (Nicotiana tabacum) seeds. It did not react with chemically deglycosylated beta-fructosidase. The anti-beta F1 serum also reacted with the bean vacuolar protein, phytohemagglutinin, but not with deglycosylated phytohemagglutinin. The anti-beta F2 antibodies recognize the beta-fructosidase polypeptide, while the beta F1 antibodies recognize glycan sidechains common to many glycoproteins. We used immunoadsorption on glycoprotein-Sepharose columns and hapten inhibition of immunoblot reactions to characterize the nature of the antigenic site. Antibody binding activity was found to be associated with Man3(Xyl)(GlcNAc)2Fuc, Man3(Xyl)(GlcNAc)2, and Man(Xyl)(GlcNAc)2 glycans, but not with Man3(GlcNAc)2. Treatment of phytohemagglutinin, a glycoprotein with a Man3(Xyl)(GlcNAc)2Fuc glycan, with Caronia lampas beta-xylosidase (after treatment with jack-bean alpha-mannosidase) greatly diminished the binding between the antibodies and phytohemagglutinin. We conclude, therefore, that the antibodies bind primarily to the xylose beta, 1 leads to 2mannose structure commonly found in the complex glycans of plant glycoproteins.