Comparison between the native glycosylated and the recombinant Cup a1 allergen: role of carbohydrates in the histamine release from basophils.
BACKGROUND
Cypress pollinosis is an important cause of respiratory allergies. Recently, the Cupressus arizonica major allergen, Cup a1, has been cloned and expressed. The native counterpart of this allergen has been purified and characterized by our group. It has been suggested that sugar moieties play a role in the in vitro IgE binding on Cupressus arizonica pollen extract.


OBJECTIVE
To characterize the immunoreactivity of the recombinant major allergen in comparison with its native counterpart. To evaluate the role of carbohydrate moieties in the IgE-mediated in vitro histamine release from basophils by using the native glycosylated Cup a1 as compared with the recombinant one.


METHODS
Recombinant Cup a1 was expressed in E. coli. IgE reactivity of Cupressaceae-allergic patients on the native as well as the recombinant molecule was investigated by immunoblotting, ELISA experiments and histamine release test from passively sensitized basophils.


RESULTS
Fourteen out of 17 Cup a1-positive sera had IgE antibodies reactive with the native molecule only and lost their reactivity-after periodate deglycosylation of the allergen. Moreover, only native molecule was capable of inducing histamine release by this group of sera. Both the recombinant and the native molecules were recognized by three out of the 17 sera and were equally capable of triggering degranulation.


CONCLUSIONS
A large number of sera reactive with the major allergen recognize carbohydrate epitopes only. IgE from these sera are able to induce histamine release from basophils and they might play a functional role in the clinical symptoms of allergy.
Iacovacci
P
P
Department of Immunology, Istituto Superiore di Sanità, Rome, Italy.
Afferni
C
C
Butteroni
C
C
Pironi
L
L
Puggioni
E M R
EM
Orlandi
A
A
Barletta
B
B
Tinghino
R
R
Ariano
R
R
Panzani
R C
RC
Di Felice
G
G
Pini
C
C
D000485:Allergens; D052179:Antigens, Plant; D001491:Basophils / Q000276:immunology; D019149:Bioreactors; D050260:Carbohydrate Metabolism; D002240:Carbohydrate Sequence; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D004926:Escherichia coli / Q000276:immunology; D006636:Histamine Release; D006801:Humans; D008969:Molecular Sequence Data; D010940:Plant Proteins; D011994:Recombinant Proteins / Q000235:genetics; D017422:Sequence Analysis, DNA
D000485:Allergens; D052179:Antigens, Plant; C419643:Cup a 1 protein, Cupressus arizonica; D010940:Plant Proteins; D011994:Recombinant Proteins