Chondroitin C lyase [4.2.2.] is unable to cleave fructosylated sequences inside the partially fructosylated Escherichia coli K4 polymer.
Chondroitin C lyase was demonstrated to be unable to act on fructosylated sequences inside a partially fructosylated polysaccharide having the chondroitin backbone structure, the Escherichia coli K4 polymer, using different analytical approaches. Chondroitin C lyase produced various unsaturated oligosaccharides by acting on an approximately 27%-fructosylated K4 polymer. The online HPLC-ESI-MS approach showed the disaccharide nature of the main species produced by chondroitinase C as DeltaHexA-GalNAc. Furthermore, the non-digested sequences inside the K4 polymer were demonstrated to be oligosaccharides bearing a fructose for each glucuronic acid unit. In fact, unsaturated fully fructosylated oligomers, from tetrasaccharide to decasaccharide (DeltaHexA(Fru)-GalNAc-[GlcA(Fru)-GalNAc](n) with n between 1 and 4), at decreasing percentages, were produced by the enzyme. These results clearly indicate that chondroitinase C cleaved the innermost glucuronic acid-N-acetylgalactosamine linkage without affecting the 1,4 glycosidic linkage between fructosylated glucuronic acid and N-acetylgalactosamine residues, confirming that the 3-O-fructosylation of the GlcA residue renders the polysaccharide resistant to the enzyme action. This novel specific activity of chondroitinase C was also useful for the production of discrete microgram amounts of fully fructosylated oligomers, from 4- to 10-mers, from E. coli K4 for possible further studies and applications.
Volpi
Nicola
N
Department of Biologia Animale, University of Modena and Reggio Emilia, Modena, Italy. volpi@unimo.it
D002240:Carbohydrate Sequence; D002808:Chondroitin Lyases / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D004187:Disaccharides / Q000032:analysis / Q000737:chemistry; D004926:Escherichia coli / Q000378:metabolism*; D005632:Fructose / Q000378:metabolism*; D008969:Molecular Sequence Data; D011108:Polymers / Q000378:metabolism*; D021241:Spectrometry, Mass, Electrospray Ionization
D004187:Disaccharides; D011108:Polymers; D005632:Fructose; D002808:Chondroitin Lyases