Glycoform analysis of Japanese cedar [Cryptomeria japonica] pollen allergen, Cry j 1

In our previous study (Y. Kimura et al., Biosci. Biotechnol. Biochem., 69, 137-144 (2005)), we found that plant complex type N-glycans harboring Lewis a epitope are linked to the mountain cedar pollen allergen Jun a 1. Jun a 1 is a glycoprotein highly homologous with Japanese cedar pollen glycoallergen, Cry j 1. Although it has been found that some plant complex type N-glycans are linked to Cry j 1, the occurrence of Lewis a epitope in the N-glycan moiety has not been proved yet. Hence, we reinvestigated the glycoform of the pollen allergen to find whether the Lewis a epitope(s) occur in the N-glycan moiety of Cry j 1. From the cedar pollen glycoallergen, the N-glycans were liberated by hydrazinolysis and the resulting sugar chains were N-acetylated and then coupled with 2-aminopyridine. Three pyridylaminated sugar chains were purified by reversed-phase HPLC and size-fractionation HPLC. The structures were analyzed by a combination of exoand endo-glycosidase digestions, sugar chain mapping, and electrospray ionization mass spectrometry (ESIMS). Structural analysis clearly indicated that Lewis a epitope (Gal beta l-3(Fuc alpha l4) GlcNAc beta 1-), instead of the Gal beta l-4(Fuc alpha l-6) GlcNAc, occurs in the N-glycans of Cry j 1.