Structure of the glycosylphosphatidylinositol membrane anchor glycan of a class-2 variant surface glycoprotein from Trypanosoma brucei

The neutral glycan fraction of the glycosylphosphatidylinositol (GPI) membrane anchor of a class-2 variant surface glycoprotein (VSG) from Trypanosoma brucei was isolated following aqueous hydrogen fluoride dephosphorylation and nitrous acid deamination of the purified glycoprotein. The neutral glycans were fractionated by high-pH anion exchange chromatography and gel-filtration and six major glycan structures were solved by a combination of one and two-dimensional NMR, composition analysis, methylation linkage analysis and electrospray-mass spectrometry. The glycans were similar to those previously described for class-1 VSGs, in that they contained the linear trimannosyl sequence Manalpha1-2Manalpha1-6Man and a complex alpha-galactose branch of up to Galalpha1-2Galalpha1-6(Galalpha1-2)Gal, but most also contained an additional galactose residue attached alpha1-2 to the non-reducing terminal mannose residue and about one-third contained an additional galactose residue attached beta1-3 to the middle mannose residue. The additional complexity of the class-2 VSG GPI glycans is discussed in terms of a biosynthetic model that explains the full range of mature GPI structures that can be expressed on different VSG classes by the same trypanosome clone.