Enzymatic properties of a ginkgo biloba endo-β-N-acetylglucosaminidase and N-Giycan structures of storage glycoproteins in the seedst

An endo-β-N-acetylglucosaminidase has been purified to homogeneity from mature seeds of Ginkgo biloba. The molecular mass of the endo-β-N-acetylglucosaminidase, named Endo-GB, was estimated to be around 63 kDa by SDS-PAGE and around 62 kDa by Hiprep S-200 chromatography, respectively. The substrate specificity has been explored with regard to the pyridylaminated N-glycans. Several high mannose-type sugar chains bearing ケ-1,2-mannosyl residue(s), Man_9-6GlcNAc₂-PA, were the most favored substrates followed by Man₅GlcNAc₂-PA and a typical hybrid-type structure (GlcNAc₁Man₅GlcNAc₂-PA) which does not bear an α-1,2-mannosyl residue. On the contrary, endo-GB could hardly hydrolyze the common core pentasaccharide of N-glycan (Man₃GlcNAc₂-PA) and the xylose-containing sugar chains (Man_4-3Xyl₁GlcNAc₂-PA, Man₃Fuc₁Xyl₁GlcNAc₂-PA) being widely distributed in plant glycoproteins. Furthermore, we analyzed the structures of N-glycans conjugated to storage glycoproteins in the mature Ginkgo seeds to see the occurrence of endogenous substrates for Endo-GB. The structural analysis showed, however, only xylose-containing type N-glycans (Man₃Fuc₁Xyl₁GlcNAc₂ (95%) and Man₃Xyl₁GlcNAc₂ (5%)), which can not be substrate for Endo-GB, predominantly occur in the storage glycoproteins.