Iwafune Mito, Kakizaki Ikuko, Nakazawa Hideaki, Nukatsuka Isoshi, Endo Masahiko, Takagaki Keiichi
Department of Biochemistry, Hirosaki University, School of Medicine, 5 Zaifu-cho, Hirosaki 036-8562, Japan.
Anal Biochem. 2004 Feb 1;325(1):35-40. doi: 10.1016/s0003-2697(03)00391-9.
Glycosaminoglycan chains were liberated from proteoglycans (bovine lung, tracheal cartilage, and cerebrum) by successive digestion with actinase and with cellulase from Aspergillus niger, which has endo-beta-xylosidase activity. The glycosaminoglycan chains were fluorescence-labeled with 2-aminopyridine after digestion with Streptomyces hyaluronidase. The resulting pyridylamino-glycosaminoglycans, including heparan sulfate, chondroitin sulfate/dermatan sulfate, and heparin, were separated by high-performance liquid chromatography. Each separated fraction was analyzed by two types of high-performance liquid chromatography: gel-filtration chromatography and anion-exchange chromatography. The correlation between molecular weight and degree of sulfation could be shown on the two-dimensional polysaccharide chain map. Use of a commonly available cellulase with endo-beta-xylosidase activity together with the two-dimensional polysaccharide chain map allows easy analysis of various glycosaminoglycan chains and comprehensive comparison among the structures. These techniques will become useful tools in the further development of glycotechnology and glycome analysis.
