Monomeric rhamnogalacturonan II (mRG-II) was isolated from red wine and the reducing-end galacturonic acid of the backbone converted to L-galactonic acid by treatment with NaBH4. The resulting product (mRG-II′ol) was treated with a cell-free extract from Penicilliumdaleae, a fungus that has been shown to produce RG-II-fragmenting glycanases. The enzymatically generated products were fractionated by size-exclusion and anion-exchange chromatographies and the quantitatively major oligosaccharide fraction isolated. This fraction contained structurally related oligosaccharides that differed only in the presence or absence of a single Kdo residue. The Kdo residue was removed by acid hydrolysis and the resulting oligosaccharide then characterized by 1- and 2D 1H NMR spectroscopy, ESMS, and by glycosyl-residue and glycosyl-linkage composition analyses. The results of these analyses provide evidence for the presence of at least two structurally related oligosaccharides in the ratio ∼6:1. The backbone of these oligosaccharides is composed of five (1→4)-linked α-d-GalpA residues and a (1→3)-linked L-galactonate. The (1→4)-linked GalpA residue adjacent to the terminal non-reducing GalpA residue of the backbone is substituted at O-2 with an apiosyl-containing side chain. β-l-Araf-(1→5)-β-d-DhapA is likely to be linked to O-3 of the GalpA residue at the non-reducing end of the backbone in the quantitatively major oligosaccharide and to O-3 of a (1→4)-linked GalpA residue in the backbone of the minor oligosaccharide. Furthermore, the results of our studies have shown that the enzymically generated aceryl acid-containing oligosaccharide contains an α-linked aceryl acid residue and a β-linked galactosyl residue. Thus, the anomeric linkages of these residues in RG-II should be revised.
NCBI PubMed ID: 10890275Publication DOI: 10.1016/s0008-6215(00)00036-7Journal NLM ID: 0043535Publisher: Elsevier
Correspondence: vidals
ensam.inra.fr
Institutions: Complex Carbohydrate Research Center, The University of Georgia, Athens, USA, Institut National de la Recherche Agronomique, Institut des Produits de la Vigne, Unité de Recherches Biopolymères et Arômes, Montpellier, France, Gist-Brocades France S.A., Seclin, France
Methods: 13C NMR, 1H NMR, NMR-2D, TLC, ESI-MS, acid hydrolysis, HPLC, extraction, CC, HR-ESI-MS, evaporation, centrifugation
Found