Fingerprinting of polysaccharides attacked by hydroxyl radicals in vitro and in the cell walls of ripening pear fruit

S C Fry, J C Dumville, Janice G. Miller

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Hydroxyl radicals ((OH)-O-.) may cause non-enzymic scission of polysaccharides in vivo, e.g. in plant cell walls and mammalian connective tissues. To provide a method for detecting the action of endogenous (OH)-O-. in vivo, we investigated the products formed when polysaccharides were treated with (OH)-O-. (generated in situ by ascorbate-H2O2-Cu2+ mixtures) followed by (NaBH4)-H-3. Treatment with (OH)-O-. increased the number of (NaBH4)-H-3-reacting groups present in citrus pectin, homogalacturonan and tamarind xyloglucan. This increase is attributed partly to the formation of glycosulose and glycosulosuronic acid residues, which are then reduced back to the original (but radioactive) sugar residues and their epimers by (NaBH4)-H-3. The glycosulose and glycosulosuronic acid residues were stable for > 16 h at 20 degreesC in ethanol or buffer (pH 4.7), but were destroyed in alkali. Driselase-digestion of the radiolabelled polysaccharides yielded characteristic patterns of H-3-products, which included galactose and galacturonate from pectin, and isoprimeverose, galactose, glucose and arabinose from xyloglucan. Pectin yielded at least eight IH-labelled anionic products, separable by electrophoresis at pH 3.5. The patterns of radioactive products form useful 'fingerprints' by which (OH)-O-.-attacked polysaccharides may be recognized. Applied to the cell walls of ripening pear (Pyrus communis) fruit, the method gave evidence for progressive (OH)-O-. radical attack on polysaccharides during the softening process.

Original languageEnglish
Pages (from-to)729-737
Number of pages9
JournalBiochemical Journal
Issue numberPart 3
Publication statusPublished - Aug 2001


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