Carbonic Anhydrase 1 Protein, Human (His, solution)

Carbonic Anhydrase 1 Protein, Human (His, solution) expresses in E. coli with a His tag at the N-terminus. Carbonic Anhydrase 1 (CA1) expression and CA1-mediated calcification are significantly associated with atherosclerosis (AS) progression^[1].

Carbonic anhydrase 1 (CA1) is a member of the carbonic anhydrase (CA) family that reversibly catalyzes the hydration of CO₂ to form HCO₃^-, which then rapidly binds to calcium ions to form calcium carbonate. CA1 was also highly expressed in human AS tissues and in rat vascular smooth muscle cells (VSMCs) with β-glycerophosphate-induced calcification^[1].

The esterase activity is determined to be greater than 24 pmol/min/μg.

Materials
Assay buffer: 12.5 mM Tris, 75 mM NaCl, pH 7.5
Test protein: Carbonic Anhydrase 1 Protein, Human (His) (HY-P7718A)
Substrate: 4-Nitrophenylacetate (4-NPA), prepared as a 20 mM stock solution in ethanol

Procedure
1. Dilute Human CA1 to 100 ng/μL in assay buffer.
2. Dilute substrate to 2 mM in assay buffer.
3. Add 50 μL of 100 ng/μL Human CA1 to the well plate, followed by 50 μL of 2 mM substrate to initiate the reaction. Perform triplicate wells.
4. Include a substrate blank containing 50 μL assay buffer and 50 μL substrate. Perform triplicate wells.
5. Read at 400 nm wavelength for 5 minutes in kinetic mode.
6. Calculate specific activity:

Human

E. coli

C-6*His

P00915 (A2-F261)

759  [NCBI]

Full Length of Mature Protein

ASPDWGYDDKNGPEQWSKLYPIANGNNQSPVDIKTSETKHDTSLKPISVSYNPATAKEIINVGHSFHVNFEDNDNRSVLKGGPFSDSYRLFQFHFHWGSTNEHGSEHTVDGVKYSAELHVAHWNSAKYSSLAEAASKADGLAVIGVLMKVGEANPKLQKVLDALQAIKTKGKRAPFTNFDPSTLLPSSLDFWTYPGSLTHPPLYESVTWIICKESISVSSEQLAQFRSLLSNVEGDNAVPMQHNNRPTQPLKGRTVRASF

25-35 kDa

• 
  ≥ 95%, as determined by reducing SDS-PAGE.

Solution

Supplied as a 0.22 μm filter solution of 12.5 mM Tris-HCl, 75 mM NaCl, pH 7.5.
Note: For SPR assay, please replace the buffer. Primary amine components (e.g., Tris, imidazole) can affect protein-coupled chips.

<1 EU/μg, determined by LAL method.

Stored at -80°C for 1 year from date of receipt. It is stable at -20°C for 3 months after opening. It is recommended to freeze aliquots at -80°C for extended storage. Avoid repeated freeze-thaw cycles.

Shipping with dry ice.

• [1]. Lin Yuan, et al. Carbonic Anhydrase 1-Mediated Calcification Is Associated With Atherosclerosis, and Methazolamide Alleviates Its Pathogenesis  [Content Brief]