Analysis of glycosylinositol phosphorylceramides expressed by the opportunistic mycopathogen Aspergillus fumigatus

Acidic glycosphingolipid components were extracted from the opportunistic mycopathogen Aspergillus fumigatus and identified as inositol phosphorylceramide and glycosylinositol phosphorylceramides (GIPCs). Using nuclear magnetic resonance sppectroscopy, mass spectrometry, and other techniques, the structures of six major components were elucidated as Ins-P-Cer (Af-0), Manp (alpha 1 -> 3) Manp(alpha 1 -> 2) Ins-P-Cer (Af-2), Manp(a1 -> 2) Manp (alpha 1 -> 3) Manp(alpha 1 -> 2) Ins-P-Cer (Af-3a), Manp(a1 -> 3) [Galf(beta 1 -> 6)] Manp(alpha 1 -> 2)-Ins-P-Cer (Af-3b), Manp (alpha 1 -> 2)-Manp(alpha 1 -> 3)[ Galf(beta 1 -> 6)] Manp(beta 1 -> 2) Ins-P-Cer (Af-4), and Manp(alpha 1 -> 3) Manp(alpha 1 -> 6) GlcpN(alpha 1 -> 2) Ins-P-Cer (Af-3c) ( where Ins 5 myo-inositol and P = phosphodiester). A minor A. fumigatus GIPC was also identified as the N-acetylated version of Af-3c (Af-3c*), which suggests that formation of the GlcN alpha 1 -> 2Ins linkage may proceed by a two-step process, similar to the GlcN alpha 1 -> 2Ins linkage in glycosylphosphatidylinositol (GPI) anchors (transfer of GlcNAc, followed by enzymatic de-N-acetylation). the glycosylinositol of Af-3b, which bears a distinctive branching Galf (b1Y6) residue, is identical to that of a GIPC isolated previously from the dimorphic mycopathogen Paracoccidioides brasiliensis (designated Pb-3), but components Af-3a and Af-4 have novel structures. Overlay immunostaining of A. fumigatus GIPCs separated on thinlayer chromatograms was used to assess their reactivity against sera from a patient with aspergillosis and against a murine monoclonal antibody (MEST-1) shown previously to react with the Galf (beta 1 -> 6) residue in Pb-3. These results are discussed in relation to pathogenicity and potential approaches to the immunodiagnosis of A. fumigatus.Univ New Hampshire, Dept Chem, Durham, NH 03824 USAUniversidade Federal de São Paulo, Escola Paulista Med, Dept Biochem, BR-04023900 São Paulo, BrazilUniv Georgia, Dept Plant Biol, Athens, GA 30602 USAUniv Georgia, Dept Biochem & Mol Biol, Athens, GA 30602 USAUniversidade Federal de São Paulo, Escola Paulista Med, Dept Biochem, BR-04023900 São Paulo, BrazilWeb of Scienc