Removes non-reducing, terminal branched fucose when linked alpha-(1-6) to GlcNAc of a GlcNAc-GlcNAc disaccharide structure that has been labeled with a fluorescent molecule. To date, the fluorescent labels ANTS, ANDSA and 2-AA have been demonstrated to function with the fucosidase. There is no cleavage of unlabeled oligosaccharides.
Sourcerec. Elizabethkingia meningosepticum (was Chryseobacterium meningosepticum)
60 µl aliquot of enzyme (30 mU) in 20 mM Tris-HCl, 25 mM NaCl,(pH 7.5).
200 µl 5x Reaction Buffer 5.0 (250 mM sodium phosphate, pH 5.0)
Specific Activity >1.5 U/mg
Activity 1.0 U/ml
Molecular weight 50,000 daltons
1. Add up to 1 nmole of labeled oligosaccharide to a tube.
2. Add de-ionized water to a total of 15 µl.
3. Add 4 µl of 5x Reaction Buffer 5.0.
4. Add 1 µl of Alpha-(1-6)-Fucosidase.
5. Incubate for 1 hour at 37˚C.
Specificity ë±(1-6) linked core fucose when covalently attached to a reporter molecule at the reducing terminal GlcNAc. The one exception is that a terminal unbranched ë±(1-3) ) or ë±(1-4) fucose is cleaved in the absence of any reporter molecule. Reporter molecules known to support cleavage are amino-napthalene disulfonic and trisulfonic acids and 2-aminobenzoic acid(2AA). However, 2-aminobenzamide(2-AB) will not support cleavage. Shorter oligosaccharides such as trimannosylchitobiose are more completely digested than longer derivatives which may require longer incubation times.
Cleavage of labeled oligosaccharides by the fucosidase is diagnostic of alpha 1-6 core fucosylation.
Specific Activity One unit of Fucosidase activity is defined as the amount of enzyme required to cleave 1 µmole of fucose from 4öÕmethylumbelliferyl-ë±-L-fucoside. in 1 minute at 37˚C and pH 5.0.
Storage Store enzyme at 4˚C.