Cross-reactivity between the major allergen from olive pollen and unrelated glycoproteins: Evidence of an epitope in the glycan moiety of the allergen (1)
The major allergen of this pollen is Ole e 1, an acidic glycoprotein of 20 kd that exhibits a high degree of polymorphism. The protein contains a single glycosylation site located at the Asn-111 residue. The glycosyl moiety of Ole e 1 was suggested to be involved in its allergenicity. Saccharides exhibiting IgE-binding capability are of great interest because of their potential involvement in the cross-reactivity between glycosylated allergens from different sources.
Ovalbumin, ascorbate oxidase, bromelain, carboxyamidomethylated HRP, and PLA2 have well-known glycan structures. Oligosaccharide content of each protein was estimated as bromelain (22 kd), one oligosaccharide unit per polypeptide chain; HRP (45 kd), seven units per polypeptide chain; ascorbate oxidase (65 kd), four units per polypeptide chain; PLA2 (16 kd), one unit per polypeptide chain; and ovalbumin (44 kd), one unit per polypeptide chain.
Ole e 1 showed two main bands: molecular weights are 18.5 kd (nonglycosylated allergen) and 20 kd (glycosylated allergen).
The main bands of the olive pollen allergen were recognized by the polyclonal antiserum. In addition, the three plant glycoproteins (ascorbate oxidase, bromelain, and HRP) were also recognized by the anti-Ole e 1 antiserum. Periodate, which sequentially removes monosaccharide residues from the nonreducing end of the sugar moiety of glycoproteins, produced completely deglycosylated forms of these glycoproteins, as deduced from the absence of binding to anti-HRP antibody. the glycoproteins immunostained share a β1→2–linked xylose on their glycan structures, which is not present in either PLA2 or ovalbumin.
The binding of the anti-Ole e 1 polyclonal serum to solid phase–fixed carboxyamidomethylated HRP, bromelain, ascorbate oxidase, ovalbumin, and PLA2 was also obtained. The relative affinities of these three interacting proteins decreased in this order: (1) HRP, (2) bromelain, (3) ascorbate oxidase. These results were corroborated by IgG-binding inhibition assays.
Native Ole e 1 (glycosylated forms: bands at 20, 22, and 40 kd) was recognized by anti-HRP but was not the chemically deglycosylated allergen. Only ascorbate oxidase, bromelain, and HRP were immunostained by IgE antibodies from patients allergic to olive pollen, and the binding was abolished after treatment with periodate.
Most of the N-linked oligosaccharides from plant glycoproteins exhibit a complex glycan with a consensus “vacuole type” structure. Ole e 1 N-linked carbohydrate could belong to this kind of glycans. Structural differences among the glycans of HRP, bromelain, and ascorbate oxidase would explain the differences observed in the relative affinities against the anti-Ole e 1 antiserum. The existence of IgE specific for carbohydrate epitopes has been demonstrated in patients with allergy. These antibodies have been suggested to be involved in cross-reactivity to a wide spectrum of biologic sources of allergens.