DEMONSTRATION OF THE PRESENCE OF ALPHA-1,6-BRANCHED SIDE-CHAINS IN THE MANNAN OF CANDIDA-STELLATOIDEA

A mild acetolysis of the mannans of Candida stellatoidea was performed after acetylation to yielded an alpha-1,6-branched mannohexaose, the presence of which had been predicted from the appearance of a specific H1-H2-correlated cross-peak in two-dimensional homonuclear Hartmann-H ahn spectroscopy. In this study, we found that the de-O-acetylation of a CO-acetyl group at the branching point, the 3,6-di-O-substituted ma nnose unit, of an acetylated oligosaccharide by sodium methoxide is si gnificantly slower than that of other acetyl groups. We could separate the 4-O-acetylated branching oligosaccharide from linear isomer using high-performance liquid chromatography. Before and after the de-O-ace tylation of the purified branching oligosaccharide, their H-1-NMR sign als were sequentially assigned by means of the nuclear Overhauser effe ct. In the sequential NMR assignment study, we showed that the alpha-1 ,6-linked mannose unit is attached to the 3-O-substituted unit based o n the presence of NOE cross-peak between H1 of the branching mannose u nit and H6 of the 3-O-substituted mannose unit. An enzyme-linked immun osorbent inhibition assay of the reactivity of factor 4 serum to C. st ellatoidea mannan by several oligosaccharides indicated that the alpha -1,6-branched oligosaccharide and the beta-1,2 linkage-containing olig osaccharides showed inhibitory activity. This result indicates that fa ctor 4 serum, as well as factor 5 and 6 sera, contains antibodies agai nst beta-1,2-linked mannose units which have been reported to particip ate in pathogenicity via cytokine production and/or adherence. From th e assignment results of H1-H2-correlated cross-peaks of oligosaccharid es and mannans, the molar ratio of the mannan side chains was proposed . In this study, we demonstrated that the epitope structure of the C. stellatoidea type I strains was the same as that of the C. albicans NI H B-792 (serotype B) strain.