Structure and characterization of the glycan moiety of L-amino-acid oxidase from the Malayan pit viper Calloselasma rhodostoma

Ophidian L-amino-acid oxidase (L-amino-acid oxygen:oxidoreductase, deaminat ing, EC 1.4.3.2) is found in the venom of many poisonous snakes (crotalids, elapids and viperids). This FAD-dependent glycoprotein has been studied fr om several snake species (e.g. Crotalus adamanteus, Crotalus atrox and Call oselasma rhodostoma) in detail with regard to the biochemical and enzymatic properties. The nature of glycosylation, however, as well as the chemical structure(s) of the attached oligosaccharide(s) are unknown. In view of the putative involvement of the glycan moiety in the biological effects of oph idian L-amino-acid oxidase, notably the apoptotic activity of the enzyme, s tructural knowledge is needed to evaluate its exact function. In this study we report on the glycosylation Of L-amino-acid oxidase from the venom of t he Malayan pit viper (Calloselasma rhodostoma). Its glycosylation is remark ably homogenous with the major oligosaccharide accounting for approximately 90% of the total sugar content. Based on detailed analysis of the isolated oligosaccharide by 2D NMR spectroscopies and MALDI-TOF mass spectrometry t he glycan is identified as a bis-sialylated, biantennary, core-fucosylated dodecasaccharide. The biological significance of this finding is discussed in light of the biological activities of the enzyme.