Acetyl esterase (sialate-O-acetylesterase) is a recombinant protein from Tannerella forsythia, ATCC 43037 strain, expressed in Escherichia coli. The enzyme removes acetyl groups attached via an O- group, mainly 9-, 8- and 7-. It can be used for monitoring of diacetylation of sialic acids on products such as erythropoietin (EPO)

Application: Acetyl esterase (sialate-O-acetylesterase) can be used to remove 9-, 8- and 7-O-acetyl groups from released sialic acids, released glycans or glycoproteins.

Source: Recombinant from Tannerella forsythia in E. Coli.

EC: 3.1.1.53

Contents:
Ludger sialate-o-acetylesterase - Kit Contents
LudgerZyme Acetyl Esterase (Supplied in PBS pH7.5 buffer containing 10 mM Tris-HCl)
LudgerZyme Acetyl Esterase RXN 10x buffer (500 mM sodium acetate pH5.5)

Number of Samples: Sufficient for up to 50 samples.

Amount of Sample: Up to 10 µg glycoprotein, up to 2.5 µg released glycans and up to 1 µg free sialic acid per digestion.

Suitable Samples: Acetyl esterase (sialate-O-acetylesterase) can act upon complex glycoprotein samples, such as erythropoietin (EPO), bovine submaxillary mucin and oral epithelial cell-bound glycans, and on N- and O-glycans released from a glycoprotein. Either fluorescently labelled or unlabelled glycans are suitable. It can also be used on released sialic acids.

Unit Definition: One unit (U) of acetyl esterase is defined as the amount of enzyme required to produce 300 µmole of 4-nitrophenol and acetate in 1 minute at 30°C in a buffer containing 50 mM Tris-HCl, 140 mM NaCl, pH 8.5, from 4-nitrophenyl acetate, a chromogenic esterase substrate.

Molecular Weight: 76.3 kDa.

Suggested usage:
1. To digest <10 µg of glycoprotein, make up a total reaction volume of 10 µL by dissolving the glycoprotein in 1 µL of LudgerZyme Acetyl Esterase RXN 10x buffer, 1 µL of LudgerZyme Acetyl Esterase and water to a final volume of 10 µL.
Use the same method for digestion of <2.5 µg released glycans or <1 µg free sialic acid.
The reaction may be scaled-up linearly to accommodate larger amounts of glycoprotein, released glycans or free sialic acids.

2. Incubate your samples in a water bath, oven or any other constant heating source at 37°C for 7 to 16 hours.
Note: Optimal incubation times and enzyme concentrations must be determined empirically for a particular substrate.
The samples are now ready for clean-up and subsequent analysis by HPLC.
Note: Optionally, the samples can be stored frozen at this point for analysis at a later date.

Storage: Store at 4°C. Protect from sources of heat and light. When stored correctly, the enzyme should be stable for 24 months from date of purchase. Exposure to ambient temperatures (20 - 26°C) over 3 days does not result in a reduction of enzymatic activity.