GALACTOSYLCERAMIDE TRANSFERASE OF TRYPANOSOMA CRUZI EPIMASTIGOTES BEAR SULFATED OLIGOSACCHARIDES

J. E. PARENTE; LANDONI M; V G DUSCHAK; A S COUTO

Potrero de los Funes

Congreso; XLVII Reunión Anual de la SAIB; 2011

Little experimental evidence exists up to date related to the mode of regulation of sphingolipids in parasites. With the aim of determining new targets for antiparasitic drugs, different enzymes of this pathway are being studied in T. cruzi. Cerebrosides are synthesized by a ceramide UDP-galactosyltransferase [CGalT] using UDP-galactose as donor and ceramide as acceptor.  In the present work, we describe the identification and partial characterization of the CGalT purified from epimastigote forms of T.  cruzi.  After different chromatographic steps a fraction containing three bands with apparent molecular weight of 70-61 kDa in SDS/PAGE was determined. This fraction presented CGalT activity. A specific CGalT antibody recognized the two higher MW bands. Knowing that CGalT is a glycoprotein, each gel band was cut and digested with PNGase F. The glycan fraction was analyzed by high performance anion exchange chromatography  (HPAE-PAD) and UV-MALDI-TOF mass spectrometry. As expected, neutral high-mannose type oligosaccharides were shown, however main signals correlate with sulfated oligosaccharides. This is the first report of the analysis of the sugar moiety of CGalT. On the other hand, these results show CGalT as the second sulfated glycoprotein described in epimastigote forms of T. cruzi.