Recombinant Human Carbonic Anhydrase XII Protein, CF

Catalog # Availability Size / Price Qty
2190-CA-010
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Recombinant Human Carbonic Anhydrase XII Protein, CF Summary

Purity
>95%, by SDS-PAGE under reducing conditions and visualized by silver stain
Endotoxin Level
<1.0 EU per 1 μg of the protein by the LAL method.
Activity
Measured by its esterase activity. The specific activity is >10 pmol/min/µg, as measured under the described conditions.
Source
Mouse myeloma cell line, NS0-derived human Carbonic Anhydrase XII/CA12 protein
Ala25-Gln291, with a C-terminal 10-His tag
Accession #
N-terminal Sequence
Analysis
Ala25
Predicted Molecular Mass
32 kDa
SDS-PAGE
38-41 kDa, reducing conditions

Product Datasheets

2190-CA

Formulation Supplied as a 0.2 μm filtered solution in Tris and NaCl.
Shipping The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.
Stability & 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.

Assay Procedure

Materials
  • Assay Buffer: 12.5 mM Tris, 75 mM NaCl, pH 7.5
  • Recombinant Human Carbonic Anhydrase XII/CA12 (rhCA12) (Catalog # 2190-CA)
  • Substrate: 4-Nitrophenyl Acetate (4-NPA) (Sigma, Catalog # N8130), 100 mM stock in acetone
  • 96-well Clear Plate (Costar, Catalog # 92592)
  • Plate reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
  1. Dilute rhCA12 to 20 ng/µL in Assay Buffer.
  2. Dilute Substrate to 2 mM in Assay Buffer.
  3. In a plate, load 50 µL of 20 ng/µL rhCA12, and start the reaction by adding 50 µL of 2 mM Substrate to wells. Include a Substrate Blank containing 50 µL Assay Buffer and 50 µL of 2 mM Substrate.
  4. Read absorbance at a wavelength of 400 nm (bottom read) in kinetic mode for 5 minutes.
  5. Calculate specific activity:

     Specific Activity (pmol/min/µg) =

Adjusted Vmax* (OD/min) x Conversion Factor** (pmol/OD)
amount of enzyme (µg)

     *Adjusted for Substrate Blank
     **Derived using calibration standard 4-Nitrophenol (Sigma, Catalog # 241326).

Per Well:
  • rhCA12: 1 µg
  • Substrate: 1 mM
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Background: Carbonic Anhydrase XII/CA12

Carbonic Anhydrase (CA) catalyzes the reversible reaction of CO2 + H2O = HCO3- + H+, which is fundamental to many processes such as respiration, renal tubular acidification and bone resorption (1). Topics in a CA meeting (6th International Conference on the CAs, June 20‑25, 2003, Slovakia) ranged from the use of CAs as markers for tumor and hypoxia in the clinic, as a nutritional supplement in milk, and as a tool for CO2 removal and mosquito control in industry. CA12 is a type I membrane enzyme highly expressed in colon, kidney, prostate, intestine and activated lymphocytes and moderately expressed in pancreas, ovary, and testis (2, 3). It is over-expressed in some renal cell cancers and in glaucoma (2, 4). Two alternatively spliced forms exist, which either contains or lacks a 11 amino acid segment just before the transmembrane domain (5). The secreted, purified recombinant human CA12 contains the residues that are common for both forms.

References
  1. Hewett-Emmett, D. and R.E. Tashian (1996) Mol. Phylogenet. Evol. 5:50.
  2. Tuereci, O. et al. (1998) Proc. Natl. Acad. Sci. USA 95:7608.
  3. Ivanov, S.V. et al. (1998) Proc. Natl. Acad. Sci. USA 95:12596.
  4. Liao, S.-Y. et al. (2003) J. Med. Genet. 40:257.
  5. Strausberg, R.L. et al. (2002) Proc. Natl. Acad. Sci. USA 99:16899.
Entrez Gene IDs
771 (Human); 76459 (Mouse)
Alternate Names
CA12; Carbonate dehydratase XII; carbonic anhydrase 12; Carbonic Anhydrase XII; carbonic anhydrase XIICAXII; carbonic dehydratase; CA-XII; EC 4.2.1.1; FLJ20151; HsT18816; Tumor antigen HOM-RCC-3.1.3

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