CreaZyme Acetyl Esterase (sialate-O-acetylesterase) Kit [For Glycan Release] (CAT#: CB-P264-K) Datasheet

Product Type
Kit
Description
Acetyl esterase (sialic acid-O-acetyl esterase) is a recombinant protein derived from Forsythiae ATCC 43037 strain and expressed in E. coli. The enzyme removes the acetyl groups attached via O-groups (mainly 9-, 8- and 7-). It can be used to monitor the diacetylation of sialic acid on products such as erythropoietin (EPO)
Features
Source: Recombinant from Tannerella forsythia in E. Coli.

EC: 3.1.1.53

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.
Applications
To remove 9-, 8- and 7-O-acetyl groups from released sialic acids, released glycans or glycoproteins. Used for characterisation of highly sialylated biotherapeutics such as EPO, FSH and blood clotting factors.
Size
50 µL; Sufficient for up to 50 samples.
Sample
Glycoproteins and glycopeptides containing N-linked glycans
Components
CreaZyme Acetyl Esterase (Supplied in PBS pH7.5 buffer containing 10 mM Tris-HCl)
CreaZyme Acetyl Esterase RXN 10x buffer (500 mM sodium acetate pH5.5)
Handling Advice
1. To digest <10 µg of glycoprotein, make up a total reaction volume of 10 µL by dissolving the glycoprotein in 1 µL of CreaZyme Acetyl Esterase RXN 10x buffer, 1 µL of CreaZyme 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
Storage: Store enzyme at 4°C.
Shelf Life
Stability: Stable at least 12 months when stored properly. Several days exposure to ambient tempertures will not reduce activity.
Research Use
For research use only. Not for human or drug use

All products and services are for Research Use Only. Do Not use in humans.

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