Preparation and application of a fluorogenic substrate for endo-beta-xylosidase.
The present paper describes a fluorometric assay for galactosaminoglycan-degrading endo-beta-xylosidase, utilizing glycosaminoglycan chains bearing a 4-methylumbelliferyl group at the reducing terminus as a substrate. This fluorogenic substrate is synthesized by human skin fibroblasts cultured in the presence of a fluorogenic xyloside, 4-methylumbelliferyl-beta-D-xyloside. The assay is based on measurement of the fluorescence of 4-methylumbelliferone, enzymatically liberated from the synthetic substrate by endo-beta-xylosidase. We examined the applicability of the assay for analysis of endo-beta-xylosidase activity.
Takagaki
K
K
Department of Biochemistry, Hirosaki University School of Medicine, Japan.
Kon
A
A
Kawasaki
H
H
Nakamura
T
T
Endo
M
M
D000818:Animals; D002478:Cells, Cultured; D004588:Electrophoresis, Cellulose Acetate / Q000379:methods; D005347:Fibroblasts / Q000201:enzymology; D006026:Glycoside Hydrolases / Q000032:analysis*; D006801:Humans; D006923:Hymecromone / Q000031:analogs & derivatives / Q000032:analysis; D007202:Indicators and Reagents; D007700:Kinetics; D008099:Liver / Q000201:enzymology; D011817:Rabbits; D012867:Skin / Q000201:enzymology; D014995:Xylosidases / Q000032:analysis* / Q000378:metabolism
D007202:Indicators and Reagents; D006923:Hymecromone; C014111:methylumbelliferyl-beta-D-xyloside; D006026:Glycoside Hydrolases; D014995:Xylosidases; C056547:endo-beta-xylosidase