Characterization ofaXylose-Specific Antiserum That Reacts withtheComplexAsparagine-Linked Glycans of Extracellular andVacuolar Glycoproteins'

Antibodies wereraised against carrot (Daucus carota) cell wall B-fructosidase thatwaseither ina native configuration (this serumiscalled anti-ftF1) orchemically deglycosylated (anti-#F2). Thetwoantisera hadcompletely different specificities when tested byimmunoblotting. Theanti-aF1 serumreacted withsfructosidase andmanyother carrot cell wallproteins aswellas withmanyproteins inextracts ofbean(Phaseolus vulgaris) cotyledons andtobacco (Nicotiana tabacum) seeds. Itdidnotreact withchemically deglycosylated al-fructosidase. Theanti-#Fl serumalsoreacted withthebeanvacuolar protein, phytohemagglutinin, butnotwithdeglycosylated phytohemagglutinin. The anti-OF2 serumreacted withbothnormal anddeglycosylated #fructosidase butnotwithother proteins. Theseresults indicate that the#F2antibodies recognize thej-fructosidase polypeptide, while thejOF, antibodies recognize glycan sidechains common tomanyglycoproteins. We usedimmunoadsorption onglycoprotein-Sepharose columns andhapteninhibition ofimmunoblotreactions tocharacterize thenatureoftheantigenic site. Antibody binding activity wasfoundtobeassociated withMan3(Xyl)(GIcNAc)2Fuc,