Characterization of beta-D-xyloside-initiated glycosaminoglycan synthesized by human skin fibroblasts in the presence of tunicamycin.
Human skin fibroblasts were incubated with a fluorogenic xyloside, 4-methylumbelliferyl-beta-D-xyloside (Xyl-MU), in the presence or absence of tunicamycin. The xyloside-initiated glycosaminoglycans (GAG-MUs) were isolated from the culture medium, and their structures characterized. When the cells were incubated with Xyl-MU in the presence of 0.2 microg ml(-1) tunicamycin, the synthesis of GAG-MU was increased about three fold, compared with the control value in the absence of tunicamycin (cells exposed to Xyl-MU alone). The structures of GAG-MUs synthesized in the presence or absence of tunicamycin were compared by HPLC analysis using gel-filtration and ion-exchange columns, enzymatic digestion, and unsaturated disaccharide composition analysis. The data indicated that cells incubated with tunicamycin produced more undersulfated and shorter GAG-MUs than cells without tynicamycin. These results suggest that tunicamycin inhibits the elongation and sulfation of glycosaminoglycan (GAG) chains and that, as a result, GAG-MUs with shorter chains and undersulfated residues, but possessing a large number of GAG chains, are synthesized in the presence of tunicamycin.
Takagaki
K
K
Department of Biochemistry, Hirosaki University School of Medicine, Japan.
Tazawa
T
T
Munakata
H
H
Nakamura
T
T
Endo
M
M
D002240:Carbohydrate Sequence; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism; D006025:Glycosaminoglycans / Q000096:biosynthesis* / Q000737:chemistry; D006027:Glycosides / Q000494:pharmacology*; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D012867:Skin / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D014415:Tunicamycin / Q000494:pharmacology*
D006025:Glycosaminoglycans; D006027:Glycosides; C010807:xylosides; D014415:Tunicamycin