RG-I is the second major component of pectic polysaccharides. Fragments of RG-I backbone composed of repeating disaccharide [^4)-a-d-GalpA-(1^2)-a-l-Rhapj can be released from commercial pectin by controlled acid hydrolysis, which allows the selective cleavage of rhamnopyranosyl linkages (Renard et al. 1998). Another method of chemical degradation of RG-I makes use of the ease of p-elimination of uronic acid esters. Heating fully methyl -esterified RG-I in mild base results in cleavage at galacturonide residues, releasing oligomers of 1-5 disaccharide repeat units terminated at the non-reducing end by 4-deoxy-P-l-t^reo-hex-4-enepyranosyluronic acid (Deng et al. 2006) (Fig. 3.2).
Enzymatic hydrolysis is a powerful tool for analysis of RG-) structure, but it is also potentially useful for the generation of oligosaccharide fragments on a preparative scale. Several enzymes are known to be capable of breaking glycosidic linkages in the RG-I backbone including RG-hydrolases and RG-lyases (Mutter et al. 1998). The former catalyse hydrolysis of GalpA-(1^2)-Rhap linkages and the latter catalyse p-elimination (as in base-catalysed fragmentation - Fig. 3.2). However, the use of enzymes for producing RG- I backbone oligosaccharides is complicated by the fact that up to 80% of the Rhap residues in RG-I are substituted. Oligosaccharide side chains, which are attached at C-4 of Rhap residues and composed predominantly of P-d-Galp and a-l-Ara/, need to be removed prior to the action of RG-hydrolase and to some extent RG-lyase. Some Galp residues remain in RG-. even after pretreatment with galactosidases; branched oligomers incorporating one or two P-d-Galp residues have been isolated (Schols et al.
1994). Although degradation techniques have been widely used for elucidation of RG- I structure, the oligosaccharide fragments that have been prepared in this way require extensive chromatographic purification and they are usually only obtained in analytical quantities. Limited availability has made comprehensive characterization of RG-I fragments difficult and their potential application as chemical or biochemical reagents is not generally practical.
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