Recombinant Human Carbonic Anhydrase XII Protein, CF
Recombinant Human Carbonic Anhydrase XII Protein, CF Summary
Ala25-Gln291, with a C-terminal 10-His tag
|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.
- 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
- Dilute rhCA12 to 20 ng/µL in Assay Buffer.
- Dilute Substrate to 2 mM in Assay Buffer.
- 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.
- Read absorbance at a wavelength of 400 nm (bottom read) in kinetic mode for 5 minutes.
- 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).
- rhCA12: 1 µg
- Substrate: 1 mM
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.
- Hewett-Emmett, D. and R.E. Tashian (1996) Mol. Phylogenet. Evol. 5:50.
- Tuereci, O. et al. (1998) Proc. Natl. Acad. Sci. USA 95:7608.
- Ivanov, S.V. et al. (1998) Proc. Natl. Acad. Sci. USA 95:12596.
- Liao, S.-Y. et al. (2003) J. Med. Genet. 40:257.
- Strausberg, R.L. et al. (2002) Proc. Natl. Acad. Sci. USA 99:16899.
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