Abstract
The complete primary structures of the major Asn-linked oligosaccharides from the type II variant surface glycoproteins (VSGs), MITat 1.2 and MITat 1.7, and the type III VSG, MITat 1.5, were determined using a combination of exo- and endoglycosidase digestions, methylation analysis, acetolysis, and 500 MHz 1H NMR spectroscopy. Each variant contained classical branched oligomannose-type and biantennary complex oligosaccharides, a proportion of the latter substituted with terminal alpha(1-3)-linked galactose residues, the first report of the presence of this epitope in Trypanosoma brucei. In addition both the type II variants contained relatively large amounts of the unusual small oligomannose-type oligosaccharides, Man4GlcNAc2 and Man3GlcNAc2, and a diverse array of novel branched poly-N-acetyllactosamine oligosaccharides, similar but not identical to those from mammalian glycoproteins. These latter structures were also partially substituted with terminal alpha(1-3)-linked galactose residues. Glycosylation in the type II variants showed site specificity in that the poly-N-acetyllactosamine and Man(9-5)GlcNAc2 oligosaccharides were located exclusively at Asn-glycosylation site 1 very close to the C terminus, whereas the Man(4-3)GlcNAc2 and biantennary complex oligosaccharides were located exclusively at site 2. This is the first report of the presence of poly-N-acetyllactosamine oligosaccharides in protozoa.
Highlights
The complete primary structuresof the major Asnlinked oligosaccharides from the type I1 variant surface glycoproteins (VSGs), MITat 1.2 and MITat 1.7, and the type I11 VSG, MITat 1.6, were determined using a combinationof exo- and endoglycosidasedigestions, methylation analysis, acetolysis, and 600 MHz ‘H NMR spectroscopy
We show further that a proportion of the biantennary complex and branched poly-N-acetyllactosamineoligosaccharides are substituted with terminal cu(l-3)-linked galactose residues, and we demonstrate a site-specific distribution of oligosaccharides in the type I1 VSGs with the exclusive location of poly-N-acetyllactosaminestructures at theC-terminal glycosylation site
We reported previously (5) the exclusive presence of classical branched oligomannose-typeoligosaccharidesat theconserved Asn-glycosylation site in the type I VSGs MITat 1.4 and MITat 1.6
Summary
The complete primary structuresof the major Asnlinked oligosaccharides from the type I1 variant surface glycoproteins (VSGs), MITat 1.2 and MITat 1.7, and the type I11 VSG, MITat 1.6, were determined using a combinationof exo- and endoglycosidasedigestions, methylation analysis, acetolysis, and 600 MHz ‘H NMR spectroscopy. There is a single variant (termed type 111)which bears no homology to cal branchedoligomannose-typeand biantennarycom- eithertype I ortype I1 variants.Bothtype I and type I1 plex oligosaccharides, a proportion of the lattersubsti- variants containone conserved Asn-glycosylationsite located tuted with terminal a(l-3)-linked galactose residues, the first reportof the presence of this epitope in Trypanosoma brucei. In addition both the typeI1 variants contained relatively largeamounts of the unusual small within the C-terminal region. Each trypanosome contains up to a thousand different VSG genes, each of which encodes an immunologicallydistinct molecule
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