Recombinant Mouse Cathepsin E Protein, CF Summary
Gln19-Pro397, with a C-terminal 10-His tag
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CF stands for Carrier Free (CF). We typically add Bovine Serum Albumin (BSA) as a carrier protein to our recombinant proteins. Adding a carrier protein enhances protein stability, increases shelf-life, and allows the recombinant protein to be stored at a more dilute concentration. The carrier free version does not contain BSA.
In general, we advise purchasing the recombinant protein with BSA for use in cell or tissue culture, or as an ELISA standard. In contrast, the carrier free protein is recommended for applications, in which the presence of BSA could interfere.
|Formulation||Supplied as a 0.2 μm filtered solution in MES 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: 50 mM Sodium Acetate, 100 mM NaCl, pH 3.5
- Recombinant Mouse Cathepsin E (rmCathepsin E) (Catalog # 1130-AS)
- Substrate: MCA-Pro-Leu-Gly-Leu-DPA-Ala-Arg-NH2 (Catalog # ES001)
- F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
- Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
- Dilute rmCathepsin E to 1.0 µg/mL in Assay Buffer.
- Incubate at room temperature for 15 minutes.
- Dilute activated rmCathepsin E to 0.04 ng/µL in Assay Buffer.
- Dilute Substrate to 40 µM in Assay Buffer.
- Load 50 µL of the 0.04 ng/µL rmCathepsin E into plate and start the reaction by adding 50 µL of 40 µM Substrate. Include a Substrate Blank containing 50 µL Assay Buffer and 50 µL of 40 µM Substrate.
- Read at excitation and emission wavelengths of 320 nm and 405 nm (top read), respectively in kinetic mode for 5 minutes.
- 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 MCA-Pro-Leu-OH (Bachem, Catalog # M-1975)
- rmCathepsin E: 0.002 μg
- Substrate: 20 µM
Background: Cathepsin E
Cathepsin E is an intracellular aspartic protease of the pepsin family (1, 2). Unlike Cathepsin D, another member of the same family and a lysosomal protease with relatively ubiquitous distribution, Cathepsin E is not a lysosomal enzyme and has a limited cell and tissue distribution. However, both Cathepsins D and E play an important role in the degradation of proteins, the generation of bioactive proteins, and antigen processing (3). Both enzymes are efficient in cleaving the Swedish mutant of amyloid precursor protein (APP) at the beta site but show almost no reactivity with the wild-type APP (4). Mouse Cathepsin E is synthesized as a precursor protein, consisting of a signal peptide (residues 1‑18), a propeptide (residues 19‑59), and a mature chain (residues 60‑397) (1).
- Tatnell, P.J. et al. (1997) FEBS Lett. 408:62.
- Kay, J. and P.J. Tatnell (2004) in Handbook of Proteolytic Enzymes (Barrett, A.J. et al., eds.), p. 33, Academic Press, San Diego.
- Tsukuba, T. et al. (2000) Mol. Cells 10:601.
- Gruninger-Leitch, F. et al. (2000) Nat. Biotechnol. 18:66.
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Fluorogenic Peptide Substrates
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