- Other Names
- Lysosomal protective protein, Protective protein Cathepsin A (PPCA), Carboxypeptidase C, Carboxypeptidase L, Protective protein for beta-galactosidase
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Cathepsin A (CTSA) is a member of S10 serine protease family and exhibits multi-enzymatic activities, such as a carboxypeptidase at an acidic condition and as a deamidase/esterase at neutral pH. CTSA is synthesized as a zymogen, a 54 kD precursor. Some evidence show that trypsin and other cathepsins (B, D, K, L) may cleave CTSA and generate mature forms of a 32kD and 20 kD disulfide-linked heterodimer in an acidic environment. CTSA forms a complex with neuraminidase (NEU1) and beta-galactosidase (GLB1) to stabilize and protect their activity against proteolysis. The deficiency in the human CTSA gene results in a lysosomal storage disorder, galactosialidosis, nephropathy, neurodegeneration, and skeletal dysplasia. In vitro and in vivo experiments have identified several CTSA substrates including peptide hormones, such as endothelin-1, bradykinin, and angiotensin I, indicating the role of CTSA in endocrine regulation. In addition, CTSA is involved in autophagy and it regulates the lysosomal-associated membrane protein type 2 (LAMP2) receptor and its degradation.Product Details
- Human Cathepsin A (CTSA), amino acids Ala29-Tyr480 (Accession#: P10629) with an N-terminal Met and a C-terminal 10-His tag, was expressed in 293E.
- Molecular Mass
- The 462 amino acid recombinant protein has a predicted molecular mass of approximately 52.78 kD. The protein migrates at approximately above 45 kD under DTT-reducing conditions and multiple bands under non-reducing conditions in SDS-PAGE. The N-terminal amino acid is Ala.
- >95%, as determined by Coomassie stained SDS-PAGE.
- 0.22 µm filtered protein solution is in 20 mM Tris, 500 mM NaCl, pH 7.5.
- Endotoxin Level
- Less than 0.1 ng per µg cytokine as determined by the LAL method.
- 10 - 100 µg sizes are bottled at 200 µg/mL.
- Storage & Handling
- Unopened vial can be stored at -20°C for three months, or at -70°C for six months. For maximum results, quick spin vial prior to opening. The protein can be aliquoted and stored from -20°C to -70°C. Stock solutions can also be prepared at 50-100 µg/mL in sterile buffer (PBS, HPBS, DPBS, or EBSS) containing carrier protein such as 0.2-1% BSA or HSA and stored in working aliquots at -20°C to -70°C. Avoid repeated freeze/thaw cycles.
- After autoactivation, Cathepsin A (CTSA) activity is determined by its ability to cleave the fluorogenic peptide substrate, Mca- Arg-Pro-Pro-Gly-Phe-Ser-Ala-Phe-Lys(DNP)-OH. The specific activity is >100 pmol/min/µg.
- Application Notes
CTSA Activity Assay Protocol
Step 1: Dilute the recombinant human CTSL to 10 µg/ml in activation buffer (25 mM MES, 5 mM DTT, pH 6.0) and incubate the protein at 25°C for 10 minutes.
Step 2: Dilute the recombinant human CTSA to 100 µg/ml in activation buffer (25 mM MES, 5 mM DTT, pH 6.0)
Step 3: Combine equal volumes of hCTSA and CTSL (final concentration: 50 µg/ml and 5 µg/ml, respectively). Incubate at 37°C for 15 minutes. Over activation may result in proteolysis and low activity.
Step 4: Dilute activated hCTSA to 2ng/µl in assay buffer (25 mM MES, 5 mM DTT, pH 5.5).
Step 5: Dilute the fluorogenic peptide substrate (Mcfa-Arg-Pro-Pro-Gly-Phe-Ser-Ala-Phe-Lys(DNP)-OH) to 20 µM using assay buffer.
Step 6: Perform the assay in a 96-well, black, flat-bottomed plate. Add 50 µl of the diluted, activated CTSA (0.1 µg/well) to a black well and start the reaction by adding 50 µl of 20 µM substrate. Assay is read on a fluorometer at excitation and emission wavelengths of 320 nm and 405 nm for five minutes.
This protein is in the latent form and needs to be activated for bioassay.
BioLegend carrier-free recombinant proteins provided in liquid format are shipped on blue-ice. Our comparison testing data indicates that when handled and stored as recommended, the liquid format has equal or better stability and shelf-life compared to commercially available lyophilized proteins after reconstitution. Our liquid proteins are validated in-house to maintain activity after shipping on blue ice and are backed by our 100% satisfaction guarantee. If you have any concerns, contact us at email@example.com.
Cathepsin A (CTSA) is ubiquitously expressed and is normally localized to lysosomes. CTSA also interacts with elastin binding protein (EBP) at the cell surface. CTSA is also identified in exosomes from pancreatic cells.
- CTSA is carboxypeptidase and deamidase that is involved in lysosomal protein degradation.
- Substance P, Neurokinin A, Eledoisin, bradykinin, angiotensin I, and oxytocin.
- Biology Area
- Apoptosis/Tumor Suppressors/Cell Death, Cell Biology, Neurodegeneration, Neuroscience, Neuroscience Cell Markers, Protein Trafficking and Clearance, Stem Cells
- Molecular Family
- Enzymes and Regulators, Lysosomal Markers
- Antigen References
1. Bonten E.J., et al. 1995. J. Biol. Chem. 270:26441-26445.
2. D'Azzo A, et al. 1982. Proc. Natl. Acad. Sci. USA. 79:4535-4539.
3. Jackman H.L., et al. 1990. J. Biol. Chem. 265:11265-11272.
4. Hiraiwa M. 1999. Cell. Mol. Life Sci. 56:894-907.
5. Shimmoto M, et al. 1993. J. Clin. Invest. 91:2393-2398.
6. Seyrantepe V, et al. 2008. Circulation 117:1973-1981.
7. Cuervo A.M., et al. 2003. EMBO J. 22:47-59.
- Gene ID
- 5476 View all products for this Gene ID
- View information about Cathepsin A on UniProt.org
- Does specific activity of a recombinant protein vary between lots?
Specific activity will vary for each lot and for the type of experiment that is done to validate it, but all passed lots will have activity within the established ED50 range for the product and we guarantee that our products will have lot-to-lot consistency. Please conduct an experiment-specific validation to find the optimal ED50 for your system.
- Have your recombinants been tested for stability?
Our testing shows that the recombinant proteins are able to withstand room temperature for a week without losing activity. In addition the recombinant proteins were also found to withstand four cycles of freeze and thaw without losing activity.
- How do you convert activity as an ED50 in ng/ml to a specific activity in Units/mg?
- Use formula Specific activity (Units/mg) = 10e6/ ED50 (ng/mL)
- How does the activity of your recombinant proteins compare to competitors?
We quality control each and every lot of recombinant protein. Not only do we check its bioactivity, but we also compare it against other commercially available recombinant proteins. We make sure each recombinant protein’s activity is at least as good as or better than the competition’s. In order to provide you with the best possible product, we ensure that our testing process is rigorous and thorough. If you’re curious and eager to make the switch to BioLegend recombinants, contact your sales representative today!
- What is the specific activity or ED50 of my recombinant protein?
The specific activity range of the protein is indicated on the product datasheets. Because the exact activity values on a per unit basis can largely fluctuate depending on a number of factors, including the nature of the assay, cell density, age of cells/passage number, culture media used, and end user technique, the specific activity is best defined as a range and we guarantee the specific activity of all our lots will be within the range indicated on the datasheet. Please note this only applies to recombinants labeled for use in bioassays. ELISA standard recombinant proteins are not recommended for bioassay usage as they are not tested for these applications.