Reversible glycosylated polypeptide is associated to Golgi membranes as a protein complex

VERÓNICA DE PINO; MARIELA GONZALEZ; LORENA NORAMBUENA; A. AZUA; ARIEL ORELLANA; SILVIA MORENO

Pinamar. Buenos Aires

Congreso; XLI Reunión de la Sociedad Argentina de Investigaciones en Bioquímica y Biología Molecular.; 2005

Sociedad Argentina de Investigaciones en Bioquímica y Biología Molecular.

REVERSIBLE GLYCOSYLATED POLYPEPTIDE IS ASSOCIATED TO GOLGI MEMBRANES AS A PROTEIN COMPLEX. De Pino, V2., Gonzalez M.1, Norambuena,L1., Azúa, A. 1, Orellana, A. 1 and Moreno, S. 2 1 Núcleo Milenio en Biología Celular Vegetal, Centro de Biotecnología Vegetal, Universidad Andres Bello, Republica 217, Santiago, Chile 2Fundación Instituto Leloir, Instituto de Investigaciones Bioquímicas, Buenos Aires-CONICET, Patricias Argentinas 435, (1405), Cap. Fed, Argentina. E-mail: smoreno@leloir.org.ar   Reversible glycosylated polypeptide is a self-glycosylating plant protein, believed to be involved in polysaccharide synthesis, however its function remains unclear. RGP protein has been localized in the trans-Golgi vesicles by immunogold labeling in pea, however little is known about the nature of its association. Previously, we described that the potato RGP protein presented different native molecular sizes with singular glycosylation properties. It is likely that the high molecular forms may be originated by an inter-glycosylation action of RGP molecules protein similarly to the mechanism proposed for oligomeric forms of glycogenin, proposed initiator of mammalian glycogen biosynthesis.  Here, we analyzed the size of the RGP forms, which are associated to Golgi vesicles. To analyze native RGP protein associated to Golgi membranes, the protein was extracted and sedimented in a 5-20% sucrose density gradient. Western blot analysis showed a distribution of RGP throughout the gradient. In addition, we analyzed the presence of RGP in protein complexes using Blue Native Gel Electrophoresis (BNGE). The results showed that RGP is part of different protein complexes associated to Golgi membranes. When these complexes were analyzed in a second dimension by SDS-PAGE followed by western blot analysis, the 38-kDa monomer of the RGP protein was found. Therefore, these results indicated that RGP associated to Golgi vesicles is part of protein complexes.