Purification of the glycoprotein allergen Ag7 from mugwort pollen by concanavalin A affinity chromatography.
Two affinity columns comprising immobilized concanavalin A (Con A), Con A-Sepharose and Con A-XP3507, were evaluated for their purifying ability for the glycoprotein allergen Ag7 from a partially purified extract of mugwort pollen. The most pronounced difference between the two columns was the nature of their nonspecific interactions; hydrophobic interactions were dominant with Con A-XP3507, whereas ionic interactions were dominant with Con A-Sepharose. Both Con A-columns were effective for purifying Ag7 with a recovery of 50% after specific elution with displacing sugars. The inclusion of 1.0 M NaCl and 20% ethylene glycol in the elution medium was useful for desorbing nonspecifically bound material, prior to specific elution of adsorbed Ag7 in the presence of the displacing sugars, alpha-methyl glucoside and alpha-methyl mannoside. The most efficient purification of Ag7 was achieved with Con A-Sepharose at room temperature rather than at 4 degrees C. Affinity chromatography with Con A-XP3507 resulted in a slightly more contaminated product (purity 54%) than with Con A-Sepharose (purity 64%).
Nilsen
B M
BM
Institute of Pharmacy, University of Oslo, Blindern, Norway.
Smestad Paulsen
B
B
Clonis
Y
Y
Mellbye
K S
KS
D000485:Allergens / Q000302:isolation & purification*; D000818:Animals; D052179:Antigens, Plant; D002846:Chromatography, Affinity / Q000379:methods; D002850:Chromatography, Gel; D003208:Concanavalin A; D006023:Glycoproteins / Q000302:isolation & purification*; D007122:Immunoelectrophoresis; D037121:Plant Lectins; D010940:Plant Proteins / Q000276:immunology / Q000302:isolation & purification*; D011058:Pollen / Q000032:analysis*; D011817:Rabbits
D000485:Allergens; D052179:Antigens, Plant; D006023:Glycoproteins; D037121:Plant Lectins; D010940:Plant Proteins; C067027:allergen Art v II protein, Artemisia vulgaris L.; D003208:Concanavalin A