Biosynthesis of O-N-acetylglucosamine-linked glycans in Trypanosoma cruzi - Characterization of the novel uridine diphospho-N-acetylglucosamine : polypeptide-N-acetylglucosaminyltransferase- catalyzing formation of N-acetylglucosamine alpha 1 -> O-threonine

• Autor(a) principal: Previato, J. O.
• Data de Publicação: 1998
• Outros Autores: Sola-Penna, M., Agrellos, O. A., Jones, C., Oeltmann, T., Travassos, Luiz Rodolpho [UNIFESP], Mendonca-Previato, L.
• Tipo de documento: Artigo
• Idioma: eng
• Título da fonte: Repositório Institucional da UNIFESP
• Texto Completo: http://dx.doi.org/10.1074/jbc.273.24.14982; http://repositorio.unifesp.br/handle/11600/25912

In this study, we have characterized the activity of a uridine diphospho-N-acetylglucosamine:polypeptide-alpha-N-acetylglucosaminyltransferase (O-alpha-GlcNAc-transferase) from Trypanosoma cruzi. the activity is present in microsomal membranes and is responsible for the addition of O-linked alpha-N-acetylglucosamine to cell surface proteins. This preparation adds N-acetylglucosamine to a synthetic peptide KPPTTTTTTTTKPP containing the consensus threonine-rich dodecapeptide encoded by T. cruzi MUC gene (Di Noia, J. M., Sanchez D. O., and Frasch, A. C. C. (1995) J. Biol. Chem. 270, 24146-24149). Incorporation of N-[H-3]acetylglucosamine is linearly dependent on incubation time and concentration of enzyme and substrate. the transferase activity has an optimal pH of 7.5-8.5, requires Mn2+, is unaffected by tunicamycin or amphomycin, and is strongly inhibited by UDP, the optimized synthetic peptide acceptor for the cytosolic O-GlcNAc-transferase (YSDSPSTST) (Haltiwanger, R. S., Holt, G. D., and Hart, G. W. (1990) J. Biol. Chem. 265, 2563-2568) is not a substrate for this enzyme. the glycosylated KPPTTTTTTTTKPP product is susceptible to base-catalyzed beta-elimination, and the presence of N-acetylglucosamine beta-linked to threonine is supported by enzymatic digestion and nuclear magnetic resonance data. These results describe a unique biosynthetic pathway for T. cruzi surface mucin-like molecules, with potential chemotherapeutic implications.