Product Description α (2-3,6,8,9) Neuraminidase Au cleaves all cleaves all non-reducing terminal sialic acid residues from complex carbohydrates and glycoproteins. The relative cleavage rates for different linkages are:α(2-6) > α(2-3) > α(2-8), α(2-9). In addition, the enzyme will cleave branched sialic acids (linked to an internal residue). This property makes it unique among neuraminidases. High concentrations of enzymes and prolonged incubation times may be required for cleaving branched residues. To cleave only non- reducing terminal α(2-3) unbranched sialic acid residues, use Neuraminidase SP, part number E-S007. α-(2-3,6,8,9) Neuraminidase is isolated from a clone of Arthrobacter ureafaciens. The enzyme has been extensively characterized using oligosaccharide standards. Contents Neuraminidase in 20 mM Tris-HCl, 25 mM NaCl, pH 7.5 Included with 20 µL and 60 µL pack sizes: 5x Reaction Buffer 250 mM sodium phosphate, pH 6.0 Molecular weight ~69,000 daltons pH optimum 6.0, active over the range 4.5-7 50 mM sodium phosphate (pH 6.0) provides the optimal buffer for enzyme activity with sialyllactose, a standard substrate. If glycosidase treatment is performed at suboptimal pH because of glycoprotein solubility or activity requirements, expect some diminution in enzyme activity. Specificity All non-reducing terminal branched and unbranched sialic acid. Formulation The enzyme is provided as a sterile-filtered solution in 20 mM Tris-HCl, 25 mM NaCl (pH 7.5). Specific Activity Assay One unit of Neuraminidase Au is defined as the amount of enzyme required to produce 1 µmole of methylumbelliferone in 1 minute at 37°C, pH 5.0 from MU-NANA (2’.-(4-methyl-umbelliferyl)-alpha-D-N acetylneuraminic acid]. Stability Stable at least 12 months when stored properly. Several days exposure to ambient temperatures will not reduce activity. Purity Neuraminidase Au is tested for contaminating protease as follows: 10 µg of denatured BSA is incubated at 37°C for 24 hours with 2 µl of enzyme. SDS-PAGE analysis of the treated BSA shows no evidence of degradation. The production host strain has been extensively tested and does not produce any detectable glycosidases. Directions for use 1. Add up to 100 µg of glycoprotein or 1 nmol of oligosaccharide to tube. 2. Add de-ionized water to a total of 14 µl. 3. Add 4 µl Reaction Buffer 6.0. 4. Add 2 µl Neuraminidase Au. 5. Incubate at 37°C for 1 hour. NOTE: longer incubation times are necessary if branched sialic acids are present. Desialylation may be monitored by SDS-PAGE if the size differential between native and de-sialylated protein is sufficient for detection. NOTE: longer incubation times are necessary if branched sialic acids are present.