Determination of the distribution of D-glucuronic acid units within the chain of pig-skin dermatan sulfate near the linkage region.

A method for analyzing the distribution of D-glucuronic acid units within the chain and near the linkage region of dermatan sulfate has been developed. The method consists of a chemical modification of the reducing terminal residue in the polysaccharide by reductive amination with excess 1,2-diaminoethane in the presence of sodium cyanoborohydride, desulfative fragmentation of the polysaccharide, labeled with 2-aminoethylamino (AEA) groups, in hot dimethyl sulfoxide containing 10% of water followed by 2,4-dinitrophenylation of the 2-aminoethylamino group, separation of the 2-(2,4-dinitrophenylamino)ethylamino labeled dermatan fragments from nonlabeled fragments on Octyl-Sepharose CL-4B gel, and determination of the uronic acid composition of the labeled fragments having various chain-length. A preparation of pig-skin dermatan sulfate (Mr 21,000, ratio of GlcA to total uronic acid, 93:500) showed an average distribution pattern of D-glucuronic acid residues near the linkage region of one N-acetylchondrosine unit in the disaccharide sequence 1-5(6) linked to the Xyl----Gal----Gal----GlcA residue, a cluster of 6-8 N-acetyldermosine units in the sequence 6(7)-12(13), and four separate N-acetylchondrosine units between the sequence adjacent to the N-acetyldermosine cluster and the sequence 23 or higher.