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Recombinant Human Acetylcholinesterase/ACHE Protein, CF

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Product Details

Summary
Reactivity HuSpecies Glossary
Applications Enzyme Activity
Format
Carrier-Free

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Recombinant Human Acetylcholinesterase/ACHE Protein, CF Summary

Details of Functionality
Measured by its ability to cleave Acetylthiocholine. The specific activity is >500 nmol/min/μg, as measured under the described conditions.
Source
Chinese Hamster Ovary cell line, CHO-derived human Acetylcholinesterase/ACHE protein
Glu32-Leu614, with a C-terminal 6-His tag
Accession #
N-terminal Sequence
Glu32
Protein/Peptide Type
Recombinant Enzymes
Gene
ACHE
Purity
>90%, by SDS-PAGE under reducing conditions and visualized by Colloidal Coomassie® Blue stain at 5 μg per lane
Endotoxin Note
<0.10 EU per 1 μg of the protein by the LAL method.

Applications/Dilutions

Dilutions
  • Enzyme Activity
Theoretical MW
65 kDa.
Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.
SDS-PAGE
60-75 kDa, reducing conditions
Publications
Read Publications using
7574-CE in the following applications:

Packaging, Storage & Formulations

Storage
Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • 6 months from date of receipt, -20 to -70 °C as supplied.
  • 3 months, -20 to -70 °C under sterile conditions after opening.
Buffer
Supplied as a 0.2 μm filtered solution in Tris and NaCl.
Purity
>90%, by SDS-PAGE under reducing conditions and visualized by Colloidal Coomassie® Blue stain at 5 μg per lane
Assay Procedure
  • Assay Buffer: 0.1 M Sodium Phosphate, 0.05% (w/v) Brij-35, pH 7.5
  • Recombinant Human Acetylcholinesterase/ACHE (rhACHE) (Catalog # 7574-CE)
  • Substrate: Acetylthiocholine (ATC) (Sigma, Catalog # A5626), 20 mM stock in DMSO
  • 5’,5’-Dithiobis(2-nitrobenzoic acid) (DTNB) (Sigma, Catalog # D-8130), 10 mM stock in DMSO
  • 96-well Clear Plate (Costar, Catalog # 92592)
  • Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
  1. Dilute rhACHE to 0.002 µg/mL in Assay Buffer.
  2. Dilute Substrate to 200 µM in Assay Buffer containing 100 µM DTNB.
  3. Load 50 µL dilute rhACHE to clear plate. Include a control containing Assay Buffer instead of rhACHE.
  4. Start reaction by adding 50 µL Substrate/DNTB mix.
  5. Read plate in kinetic mode for 5 minutes at an absorbance of 405 nm.
  6. Calculate specific activity:

     Specific Activity (nmol/min/µg) =

Adjusted Vmax* (OD/min) x well volume (L) x 109 nmol/mol
ext. coeff** (M-1cm-1) x path corr.*** (cm) x amount of enzyme (µg)

     *Adjusted for Substrate Blank 
     **Using the extinction coefficient 13260 M-1cm-1 
     ***Using the path correction 0.32 cm
     Note: the output of many spectrophotometers is in mOD Per Well:
  • rhACHE: 0.0001 µg
  • DTNB: 50 µM
  • ATC: 100 µM

Notes

Coomassie is a registered trademark of Imperial Chemical Industries Ltd.

This product is produced by and ships from R&D Systems, Inc., a Bio-Techne brand.

Alternate Names for Recombinant Human Acetylcholinesterase/ACHE Protein, CF

  • acetylcholinesterase (Yt blood group)
  • Acetylcholinesterase
  • ACHE
  • apoptosis-related acetylcholinesterase
  • ARACHE
  • EC 3.1.1
  • EC 3.1.1.7
  • N-ACHE
  • Yt blood group
  • YT

Background

The classical role of ACHE is to terminate cholinergic neurotransmission by hydrolysis of acetylcholine (ACH) (1). ACHE is thought to be involved in the pathology of Alzheimer's disease (AD) by accelerating the assembly of A beta peptides into fibrillar species through forming complexes with A beta via the peripheral anionic site on ACHE. ACHE inhibitors have been used to delay symptoms of AD patients by virtue of their ability to enhance ACH availability, as well as reduce amyloidogenesis and subsequent neurotoxicity (2). Its involvement in the cholinergic anti-inflammatory pathway connects ACHE with a possible marker of low-grade systemic inflammation in obesity, hypertension, coronary heart disease, and AD (3). Alternative splicing produces three isoforms: an amphipathic form that exists as both monomeric and multimeric forms, a soluble monomeric form lacking the cysteine residue near the C-terminus, and a GPI-anchored dimeric form found in the membranes of erythrocytes (1). The recombinant human ACHE (rhACHE) was expressed as the amphipathic form that consists of soluble monomer and multimeric forms.

  1. Grisaru, D. et al. (1999) Eur. J. Biochem, 264:672.
  2. Campbell, V. A. and Gowran, A. (2007) Br. J. Pharm. 152:655.
  3. Das, U. N. (2007) Med. Sci. Monit. 13:RA214.

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Publications for Acetylcholinesterase/ACHE (7574-CE)(3)

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Bioinformatics

Gene Symbol ACHE
Uniprot