Characteristics of xyloglucan after attack by hydroxyl radicals.
It has been proposed that plant cell-wall polysaccharides are subject in vivo to non-enzymic scission mediated by hydroxyl radicals (-*OH). In the present study, xyloglucan was subjected in vitro to partial, non-enzymic scission by treatment with ascorbate plus H(2)O(2), which together generate -*OH. The partially degraded xyloglucan appeared to contain ester bonds within the backbone, as indicated by an irreversible decrease in viscosity upon alkaline hydrolysis. Aldehyde and/or ketone groups were also introduced into the polysaccharide by -*OH-attack, as indicated by staining with aniline hydrogen-phthalate and by reaction with NaB(3)H(4). The introduction of ester and oxo groups supports the proposed sequence of reactions: (a) -*OH-mediated H-abstraction to produce a carbon-centred carbohydrate radical; (b) reaction of the latter with O(2); and (c) elimination of a hydroperoxyl radical (HO(2)*-). When the partially degraded xyloglucan was reduced with NaB(3)H(4) followed by acid hydrolysis, several 3H-aldoses were detected ([3H]galactose, [3H]xylose, [3H]glucose, [3H]ribose and probably [3H]mannose), in addition to unidentified 3H-products (probably including anhydroaldoses). 3H-Alditols were undetectable, showing that few or no conventional reducing termini were introduced. Digestion of the NaB(3)H(4)-reduced, partially degraded xyloglucan with Driselase released 25 times more [3H]Xyl-alpha-(1-->6)-Glc than Xyl-alpha-(1-->6)-[3H]Glc, suggesting that the xylose side-chains of the xyloglucan had been more heavily attacked by -*OH than the glucose residues of the backbone. The radioactive xyloglucan was readily digested by cellulase, yielding 3H-products in the hepta- to nonasaccharide range. A fingerprinting strategy for identifying -*OH-attacked xyloglucan in plant cell walls is proposed.
Miller
J G
JG
Edinburgh Cell Wall Group, Institute of Cell and Molecular Biology, The University of Edinburgh, Daniel Rutherford Building, The King's Buildings, Mayfield Road, EH9 3JH, Edinburgh, UK.
Fry
S C
SC
D001205:Ascorbic Acid; D001894:Borohydrides; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D007887:Fabaceae / Q000737:chemistry; D005690:Galactose / Q000032:analysis; D005936:Glucans; D005947:Glucose / Q000032:analysis; D006861:Hydrogen Peroxide; D017665:Hydroxyl Radical; D008358:Mannose / Q000032:analysis; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D010946:Plants, Medicinal; D011134:Polysaccharides / Q000737:chemistry*; D012266:Ribose / Q000032:analysis; D012639:Seeds / Q000737:chemistry; D014316:Tritium; D014990:Xylans; D014994:Xylose / Q000032:analysis
D001894:Borohydrides; D005936:Glucans; D009838:Oligodeoxyribonucleotides; D011134:Polysaccharides; D014990:Xylans; D014316:Tritium; D017665:Hydroxyl Radical; C029353:xyloglucan; D012266:Ribose; C025364:sodium borohydride; D014994:Xylose; D006861:Hydrogen Peroxide; D005947:Glucose; D008358:Mannose; D001205:Ascorbic Acid; D005690:Galactose