GENERATION AND STRUCTURAL CHARACTERIZATION OF A RANGE OF UNMODIFIED CHONDROITIN SULFATE OLIGOSACCHARIDE FRAGMENTS

Chondroitin sulfate C has been used to demonstrate an approach of gene rating a range of unmodified glycosaminoglycan oligosaccharide fragmen ts. This involves cleavage by oxymercuration treatment of the nonreduc ing terminal 4,5-unsaturated uronic acid (DeltaUA) residues from the f ragments produced by enzymatic digestion of chondroitin sulfate with c hondroitinase ABC. Carrying out the reaction on the unfractionated dig estion mixture produces a range of mono- to tridecasaccharides, the co mpositions of which were established by liquid secondary ion mass spec trometry (LSIMS) and their chromatographic patterns compared with olig osaccharides in the untreated digest. Ten of the main sequences, tri- to octasaccharides, isolated by HPLC from the treated and untreated di gests were fully characterized by a combination of LSIMS and(1)H NMR. Of these, 6 are homologs of the series with structures aUA1-[3GalNAc(6 S)Beta1-4GlcABeta1](n)-3Gal-NAc(6S) and [GalNAc(6S)Beta1-4GlcABeta1](n )-3GalNAc(6S), where n=1-3. The other 4 sequences, aUA1-[3Gal-NAc(6S)B eta1-4GlcABeta1](n)-3GalNAc(4S) and [GalNAc(6S)Beta1-4GlcABeta1](n)-3G alNAc(4S), where n=1 and 2, contain the alternative 4-sulfated GalNAc at the reducing terminal. These results establish that oligosaccharide s generated by oxymercuration treatment retain their integrity and onl y lack the terminal DeltaUA residue.