Recombinant Human AMCase/CHIA Protein, CF

• Reactivity: Hu
• Applications: Enzyme Activity
• Format: Carrier-Free

Recombinant Human AMCase/CHIA Protein, CF Summary

• Details of Functionality: Measured by its ability to cleave 4-Methylumbelliferyl-beta -D-N,N’,N”-triacetylchitotriose (4-MU-TACT). The specific activity is >125 pmol/min/μg, as measured under the described conditions.
• Source: Human embryonic kidney cell, HEK293-derived human AMCase/CHIA protein
  Tyr22-Pro408, with a C-terminal 6-His tag
• Accession #: Q9BZP6-1
• N-terminal Sequence: Tyr22
• Protein/Peptide Type: Recombinant Proteins
• Purity: >95%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.
• Endotoxin Note: <0.10 EU per 1 μg of the protein by the LAL method.

Applications/Dilutions

• Dilutions:
  • Enzyme Activity
• Theoretical MW: 44 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: 39-47 kDa, reducing conditions

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: >95%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.
• Assay Procedure:
  • Assay Buffer: 0.1 M MES, pH 5.0
  • Recombinant Human AMCase/CHIA (rhCHIA) (Catalog # 9859-GH)
  • Substrate: 4-Methylumbelliferyl-beta -D-N, N', N''-triacetylchitotrioside (Sigma, Catalog # M5639), 5 mM stock in DMSO
  • F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
  • Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
  1. Dilute rhCHIA to 4 ng/μL in Assay Buffer.
  2. Dilute Substrate to 400 μM in Assay Buffer.
  3. Load into a plate 50 μL of 4 ng/μL rhCHIA, and start the reaction by adding 50 μL of 400 μM Substrate. Include Substrate Blank containing 50 μL of Assay Buffer and 50 μL of 400 μM Substrate.
  4. Read plate at (excitation and emission) of 365 nm and 445 nm, respectively, (top read) in kinetic mode for 5 minutes.
  5. Calculate specific activity:

  Specific Activity (pmol/min/µg) = (Adjusted Vmax* (RFU/min) x Conversion Factor** (pmol/RFU)) / (amount of enzyme (µg))

  *Adjusted for Substrate Blank
  **Derived using calibration standard 4-Methylumbelliferone (Sigma, Catalog # M1381)

  Per Well:
  • rhCHIA: 0.2 μg
  • Substrate: 200 μM

Notes

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

Alternate Names for Recombinant Human AMCase/CHIA Protein, CF

• acidic mammalian chitinase
• AMCase
• CHIA
• CHIT2
• CHIT2DKFZp313J1722
• Chitinase, Acidic
• EC 3.2.1.14
• Lung-Specific Protein TSA1902
• TSA1902AMCase

Background

Human acidic mammalian chitinase (AMCase), or CHIA, degrades chitin and chitotriose, therefore participates in the defense against nematodes, fungi and other pathogens (1, 2, 3). It is one of the important proteins involved in T-helper cell type 2 (Th2)-mediated inflammation and has been implicated in asthma and allergic diseases (4). It protects lung epithelial cells against apoptosis possibly via promoting phosphorylation of AKT1 and stimulating chemokine production (5, 6). Inhibition of AMCase results in decreased airway inflammation and airway hyper-responsiveness in a mouse asthma model. Its function in the inflammatory response and in protecting cells against apoptosis is inhibited by allosamidin, suggesting that the function of this protein depends on carbohydrate binding (7).

1. Boot, R.G. et al. (2001) J. Biol. Chem. 276:6770.
2. Seibold, M.A. et al. (2009) J. Biol. Chem. 284:19650.
3. Chou, Y.T. et al. (2006) Biochemistry 45:4444.
4. Zhu, Z, et al. (2004) Science 304:1678.
5. Hartl, D. et al. (2008) J. Biol. Chem. 283:33472.
6. Hartl, D. et al. (2009) J. Immunol. 182:5098.
7. Ober, C. and Chupp, G.L. (2009) Curr. Opin. Allergy Clin. Immunol. 9:401.

Publications for AMCase/CHIA (9859-GH)(1)

Publication using 9859-GH

• Wu, Y;Adeniyi-Ipadeola, G;Adkins-Threats, M;Seasock, M;Suarez-Reyes, C;Fujiwara, R;Bottazzi, ME;Song, L;Mills, JC;Weatherhead, JE; Host gastric corpus microenvironment facilitates Ascaris suum larval hatching and infection in a murine model PLoS neglected tropical diseases 2024-02-01 [PMID: 38324590] (Bioassay, Ascaris Suum)

Bioinformatics

• Uniprot:
  • Q9BZP6-1()