TY - JOUR
T1 - Carbohydrate moieties on the in vitro immunoreactivity of soy β-conglycinin
AU - Amigo-Benavent, Miryam
AU - Athanasopoulos, Vasileios I.
AU - Ferranti, Pasquale
AU - Villamiel, Mar
AU - del Castillo, M. Dolores
PY - 2009/8
Y1 - 2009/8
N2 - β-Conglycinin is a functional glycoprotein and one of the most important soybean allergens. The aim of the present research was to investigate the role of the N-glycans moieties of β-conglycinin on its in vitro immunoreactivity. The soy allergen was obtained by isoelectric precipitation from commercial soy protein isolate and was enzymatically deglycosylated by PNGase F (Peptide N-Glycosidase F EC 3.5.1.52). In order to optimize deglycosylation conditions different reaction times and allergen concentrations were tested. The extent of deglycosylation was estimated by SDS-PAGE, CZE, RP-HPLC, and MALDI-TOF MS analyses, which provided information related to changes in protein structure. The antigenicity of both native β-conglycinin and its deglycosylated form was evaluated by western-blotting and indirect ELISA employing polyclonal rabbit anti-soybean sera and horseradish peroxidase-labeled goat anti-rabbit IgG while the in vitro allergenicity was assessed by means of indirect competitive inhibition ELISA employing human sera (IgE) of soy allergics. β-Conglycinin was effectively deglycosylated by PNGase F. Data on immunological tests suggested that glycosyl moieties forming this glycoprotein might be involved in its immunoreactivity.
AB - β-Conglycinin is a functional glycoprotein and one of the most important soybean allergens. The aim of the present research was to investigate the role of the N-glycans moieties of β-conglycinin on its in vitro immunoreactivity. The soy allergen was obtained by isoelectric precipitation from commercial soy protein isolate and was enzymatically deglycosylated by PNGase F (Peptide N-Glycosidase F EC 3.5.1.52). In order to optimize deglycosylation conditions different reaction times and allergen concentrations were tested. The extent of deglycosylation was estimated by SDS-PAGE, CZE, RP-HPLC, and MALDI-TOF MS analyses, which provided information related to changes in protein structure. The antigenicity of both native β-conglycinin and its deglycosylated form was evaluated by western-blotting and indirect ELISA employing polyclonal rabbit anti-soybean sera and horseradish peroxidase-labeled goat anti-rabbit IgG while the in vitro allergenicity was assessed by means of indirect competitive inhibition ELISA employing human sera (IgE) of soy allergics. β-Conglycinin was effectively deglycosylated by PNGase F. Data on immunological tests suggested that glycosyl moieties forming this glycoprotein might be involved in its immunoreactivity.
KW - β-Conglycinin
KW - Deglycosylation
KW - Immunoreactivity
KW - PNGase F
KW - Soy (Glycine max)
UR - http://www.scopus.com/inward/record.url?scp=67349117720&partnerID=8YFLogxK
U2 - 10.1016/j.foodres.2009.03.003
DO - 10.1016/j.foodres.2009.03.003
M3 - Article
AN - SCOPUS:67349117720
SN - 0963-9969
VL - 42
SP - 819
EP - 825
JO - Food Research International
JF - Food Research International
IS - 7
ER -