,17 as well as a polysaccharide check details component in Chlorella vulgaris.18 The α-glucan and rhamnomannans were obtained from P. boydii by extraction with hot 2% aqueous potassium hydroxide at 100 °C followed by fractionation on a Superdex 200 column (Fig. 4).11,13,14 The chemical structure of the glucan P. boydii was determined, using a combination of techniques including gas chromatography, 1H TOCSY, 1H and 13C NMR spectroscopy and methylation analysis.11 Its structure resembles
glycogen, since it consisted of (14)-linked α-D-Glcp substituted at O-6 with α-D-Glcp units (Fig. 5a and b). Identification of rhamnomannan was by mono-dimensional NMR (1H and 13C) and bi-dimensional COSY, TOCSY and HSQC analyses. The NMR data of the rhamnomannan showed anomeric signals with δ 97.9/4.981, 101.0/4.967,
102.2/5.228 and 103.9/5.060, typical of non-reducing terminal α-Rhap, and 3,6-di-O-substituted 2-O- and 3-O-substituted α-Manp units, respectively. That at δ 79.9/4.127 confirmed the presence of 3-O-substituted α-Manp units.13,14 Polysaccharides and peptidopolysaccharides are especially relevant for the architecture of the Scedosporium/P. boydii cell wall, but Luminespib several of them are immunologically active with great potential as regulators of pathogenesis and the immune response of the host. In addition, some of these molecules can be specifically recognised by antibodies from the sera of patients, suggesting that they could also be useful in the diagnosis of fungal infections. The structures of PRM-Sp of S. prolificans, as already mentioned, differed from those present in the PRM of P. boydii, which contained a higher proportion of (13)-, but no (12)-linked α-Rhap units. These structural differences in the carbohydrate portion suggest that related infections caused by P. boydii and S. prolificans
would be distinguishable by ELISA using hyperimmune sera against their component PRMs (Fig. 6a and b). Rhamnose-containing structures appear to Isotretinoin be the immunodominant epitopes in the rhamnomannans of P. boydii,7,8S. prolificans, S. schenckii and Ceratocystis stenoceras,15 particularly if they are present as (13)-linked α-Rhap side-chain units.19 Antibodies recognising this structure may, therefore, recognise both the N-linked high molecular weight polysaccharides and the O-linked oligosaccharides in the glycocomplexes. The O-glycosidically terminated oligosaccharides may account for a significant part of the PRM antigenicity, since de-O-glycosylation decreased its activity by 70–80%.8 Similar results were obtained with the peptidogalactomannan from Aspergillus fumigatus20 and PRM from S. schenckii.15 The immunodominance of the O-linked oligosaccharide chains was evaluated testing their ability to inhibit reactivity between the PRM and anti-P. boydii rabbit antiserum in an enzyme-linked immunosorbent assay (ELISA) hapten system.