Structural characterization of the pectic polysaccharide rhamnogalacturonan II: evidence for the backbone location of the aceric acid-containing oligoglycosyl side chain.
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 Penicillium daleae, 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 approximately 6:1. The backbone of these oligosaccharides is composed of five (1-->4)-linked alpha-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. Beta3-L-Araf-(1-->5)-beta-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 alpha-linked aceryl acid residue and a beta-linked galactosyl residue. Thus, the anomeric linkages of these residues in RG-II should be revised.
Vidal
S
S
Institut National de la Recherche Agronomique, Institut des Produits de la Vigne, Unité de Recherches Biopolymères et Arômes, Montpellier, France. vidals@ensam.inra.fr
Doco
T
T
Williams
P
P
Pellerin
P
P
York
W S
WS
O'Neill
M A
MA
Glushka
J
J
Darvill
A G
AG
Albersheim
P
P
D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002473:Cell Wall / Q000737:chemistry; D002474:Cell-Free System; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009005:Monosaccharides / Q000737:chemistry / Q000378:metabolism; D009844:Oligosaccharides / Q000737:chemistry* / Q000378:metabolism; D010368:Pectins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D010407:Penicillium / Q000201:enzymology*; D017421:Sequence Analysis; D014920:Wine
D009005:Monosaccharides; D009844:Oligosaccharides; C042492:rhamnogalacturonan II; D010368:Pectins