Structural analysis of the N-glycans of the major cysteine proteinase of Trypanosoma cruzi: Identification of sulfated high-mannose type oligosaccharides

Mariana Barboza Gardner, Vilma G. Duschak, Yuko Fukuyama, Hiroshi Nonami, Rosa Erra-Balsells, Juan J. Cazzulo, Alicia S. Couto

Research output: Contribution to journalArticlepeer-review

45 Scopus citations

Abstract

Trypanosoma cruzi, the parasitic protozoan that causes Chagas disease, contains a major cysteine proteinase, cruzipain. This lysosomal enzyme bears an unusual C-terminal extension that contains a number of post-translational modifications, and most antibodies in natural and experimental infections are directed against it. In this report we took advantage of UV-MALDI-TOF mass spectrometry in conjunction with peptide N-glycosidase F deglycosylation and high performance anion exchange chromatography analysis to address the structure of the N-linked oligosaccharides present in this domain. The UV-MALDI-TOF MS analysis in the negative-ion mode, using nor-harmane as matrix, allowed us to determine a new striking feature in cruzipain: sulfated high-mannose type oligosaccharides. Sulfated GlcNAc2Man3 to GlcNAc 2Man9 species were identified. In accordance, after chemical or enzymatic desulfation, the corresponding signals disappeared. In addition, by UV-MALDI-TOF MS analysis (a) a main population of high-mannose type oligosaccharides was shown in the positive-ion mode, (b) lactosaminic glycans were also identified, among them, structures corresponding to monosialylated species were detected, and (c) as an interesting fact a fucosylated oligosaccharide was also detected. The presence of the deoxy sugar was further confirmed by high performance anion exchange chromatography. In conclusion, the total number of oligosaccharides occurring in cruzipain was shown to be much higher than previous estimates. This constitutes the first report on the presence of sulfated glycoproteins in Trypanosomatids.

Original languageEnglish (US)
Pages (from-to)3803-3815
Number of pages13
JournalFEBS Journal
Volume272
Issue number15
DOIs
StatePublished - Aug 1 2005
Externally publishedYes

Keywords

  • Cruzipain
  • Nor-harmane
  • Sulfated oligosaccharides
  • Trypanosoma cruzi
  • UV-MALDI-TOF MS

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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