Glucuronoxylomannan of Cryptococcus neoformans serotype B: structural analysis by gas-liquid chromatography-mass spectrometry and 13C-nuclear magnetic resonance spectroscopy

Abstract

The major extracellular polysaccharide (glucuronoxylomannan, GXM) from six strains of Cryptococcus neoformans serotype B was characterized by gas-liquid chromatography (g.l.c.), g.l.c.-mass spectrometry (g.l.c.-m.s.), and nuclear magnetic resonance (n.m.r.) spectroscopy. Ultrasonic irradiation (u.i.) was used to reduce the mol.wt. of native GXM from 9.75 x 10(5) to 1.15 x 10(5) without apparent change in its composition (GXM-S). The Xylp:Manp:GlcpA molar ratio of the GXM and GXM-S from the six strains of C. neoformans serotype B is approximately 3.5:3.0:0.6. GXM-S was O-deacetylated (GXM-D) by treatment with NH4OH. The 13C-n.m.r. analysis of GXM-D gave spectra that served as characteristic fingerprints of the structure and also facilitated the assignment of the anomeric carbon resonances to specific structural moieties present in GXM-D. The GXM-D from each serotype B strain was found to be similar by 13C-n.m.r. spectroscopy. The structure contains a linear (1----3)-alpha-D-Manp backbone substituted with 2-O-beta-GlcpA and 2-O-beta-Xylp. beta-Xylp is also O-4 linked to the Manp substituted with GlcpA. In addition, a model for the disposition of the Xylp and GlcpA side chain substituents along the mannopyranan backbone is proposed, based upon results from the combination of g.l.c.-m.s. and 13C-n.m.r. spectroscopy.