A fluorescence-quenched chitopentaose for the study of endo-chitinases and chitobiosidases.

摘要: A new fluorogenic substrate displaying intramolecular fluorescence energy transfer (FRET) has been synthetized from NI,NII,NIII, NIV-tetra-acetyl-chitopentaose. Two molecules, a fluorophore (5-(2-aminoethyl) amino-1-naphtalene-sulfonic acid; EDANS) and quenching group (dimethylaminophenylazophenyl; DAB) were chemically introduced on to the chitopentaose, one at each end. Among eight enzymes tested, only endo-chitinase chitobiosidase activities could be specifically assayed by monitoring variation of after enzymatic hydrolysis this substrate. Chitobiases N-acetyl-beta-glucosaminidases are not active compound, presence bulky chromogenic 2 position nonreducing end subtrate preventing binding thus these two exo-enzymes. The observation that chitobiosidases able hydrolyse chitooligosaccharide functionalized both extremities demonstrates possibility an endo-action for class chitinases, which generally classified as This should prove very useful detection convenient assay chitinolytic nanomolar concentrations.