Identification of oligomeric domains within dermatan sulfate chains using differential enzymic treatments, derivatization with 2-aminoacridone and capillary electrophoresis.
Galactosaminoglycans, i.e. dermatan sulfate (DS) and chondroitin sulfate, are linear heteropolysaccharides consisting of repeating disaccharide units of L-iduronic acid (L-IdoA) or D-glucuronic acid (D-GlcA) residues linked to N-acetyl-galactosamine. High-performance capillary electrophoresis (HPCE or CE) has been successfully used for determining the disaccharide composition of glycosaminoglycans. However, only limited information is available on how to identify oligomeric domains rich in D-GlcA or L-IdoA. The aim of this study was therefore to develop a rapid and accurate CE procedure by which such oligosaccharides can be determined together with the variously sulfated disaccharides. Isolated dermatan sulfates of human origin were separately digested with chondroitinases ABC, AC and B and the enzymic products were derivatized with 2-aminoacridone. CE analysis of these products was performed using a phosphate buffer, pH 3.0, and reversed polarity at 30 kV. The derivatization enabled their detection with laser-induced fluorescence (LIF) and UV at 260 nm at much higher sensitivity than the detection of nonderivatized delta-saccharides at 232 nm and therefore components undetectable at 232 nm were nicely detected after derivatization. Except for delta-disaccharides, altogether five distinct oligosaccharides with differences in charge density were identified. Depending on the lyase that produced these oligomers, information on the presence of L-IdoA- or D-GlcA-containing domains within the DS chain and the sulfation pattern of these oligomeric domains was obtained. This CE method could also be useful in studying the functional oligomeric domains in galactosaminoglycan chains.
Mitropoulou
T N
TN
Department of Chemistry, University of Patras, Greece.
Lamari
F
F
Syrokou
A
A
Hjerpe
A
A
Karamanos
N K
NK
D000609:Aminoacridines; D001426:Bacterial Proteins / Q000378:metabolism; D019765:Chondroitin ABC Lyase / Q000378:metabolism; D002808:Chondroitin Lyases / Q000378:metabolism; D002810:Chondroitinases and Chondroitin Lyases / Q000378:metabolism; D003871:Dermatan Sulfate / Q000737:chemistry*; D004187:Disaccharides / Q000032:analysis; D019075:Electrophoresis, Capillary / Q000379:methods*; D005260:Female; D020723:Glucuronic Acid / Q000032:analysis; D006801:Humans; D007067:Iduronic Acid / Q000032:analysis; D007889:Leiomyoma / Q000737:chemistry; D009215:Myometrium / Q000737:chemistry; D009844:Oligosaccharides / Q000032:analysis; D011134:Polysaccharides / Q000737:chemistry; D017434:Protein Structure, Tertiary; D013379:Substrate Specificity; D014594:Uterine Neoplasms / Q000737:chemistry
D000609:Aminoacridines; D001426:Bacterial Proteins; D004187:Disaccharides; D009844:Oligosaccharides; D011134:Polysaccharides; D003871:Dermatan Sulfate; C072515:2-aminoacridone; D007067:Iduronic Acid; D020723:Glucuronic Acid; C007815:galactosaminoglycan; D002808:Chondroitin Lyases; D002810:Chondroitinases and Chondroitin Lyases; C446037:chondroitinase B; D019765:Chondroitin ABC Lyase