Recombinant Human Proprotein Convertase 1/PCSK1 Protein, CF
Recombinant Human Proprotein Convertase 1/PCSK1 Protein, CF Summary
Product Specifications
Ser111-Arg617, with a C-terminal 10-His tag
Analysis
Product Datasheets
Carrier Free
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.
2810-SE
| Formulation | Supplied as a 0.2 μm filtered solution in Tris, NaCl and Glycerol. |
| Shipping | The product is shipped with dry ice or equivalent. Upon receipt, store it immediately at the temperature recommended below. |
| Stability & Storage: | Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
|
Assay Procedure
- Assay Buffer: 25 mM MES, 5 mM CaCl2, 1% (w/v) Brij-35, pH 6.0.
- Recombinant Human Proprotein Convertase 1/PCSK1 (rhPCSK1) (Catalog # 2810-SE)
- Substrate: L-PyroGlu-Arg-Thr-Lys-Arg-AMC (pERTKR-AMC) (Catalog # ES013)
- F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
- Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
- Dilute rhPCSK1 to 4 µg/mL in Assay Buffer.
- Dilute Substrate to 200 µM in Assay Buffer.
- Load 50 µL of 4 µg/mL of rhPCSK1 into a plate, and start the reaction by adding 50 µL of 200 µM Substrate. Include a Substrate Blank containing 50 µL of Assay Buffer and 50 µL of Substrate.
- Incubate at 37 °C for 1 hour.
- Read at excitation and emission wavelengths of 380 nm and 460 nm (top read), respectively in endpoint mode.
- Calculate specific activity:
|
Specific Activity (pmol/min/µg) = |
Adjusted Fluorescence* (RFU) x Conversion Factor** (pmol/RFU) |
| Incubation time (min) x amount of enzyme (µg) |
*Adjusted for Substrate Blank
**Derived using calibration standard 7-Amino, 4-Methyl Coumarin (AMC) (Sigma, Catalog # A9891).
- rhPCSK1: 0.200 µg
- Substrate: 100 µM
Reconstitution Calculator
Background: Proprotein Convertase 1/PCSK1
The human PCSK1 gene encodes Proprotein Convertase 1 (PC1), which is also known as PC3 (1, 2). As a serine protease of the furin/PC family, the deduced amino acid sequence of human PC1 consists of a signal peptide (residues 1 to 27), a pro peptide (residue 28 to 110), and a mature chain (residues 111 to 753). Autocatalytic processing results in the removal of the pro peptide as well as a region from the C-terminus, which leads to the production of the most active 66 kDa form (3). The purified rhPC1 consists of residues 111 to 617, is active in the assay described above and most likely corresponds to the 66 kDa form. Found almost exclusively in endocrine and neural cells and stored as the 66 kDa form in dense-core secretory vesicles, PC1 is capable of processing several endocrine and neural prohormones, such as opiomelanocortin, somatostatin, enkephalin, insulin, dynorphin and thyrotropin-releasing hormone (3). Indeed, obesity and impaired prohormone processing have been associated with mutations in the human PCSK1 gene (4).
- Creemers, J.W. et al. (1992) FEBS Lett. 300:82.
- Seidah, N.G. et al. (1992) DNA Cell Biol. 11:283.
- Seidah, N.G. and M. Chretien (2004) in Handbook of Proteolytic Enzymes (ed. Barrett, et al.) p. 1861. Academic Press, San Diego.
- Jackson, R.S. et al. (1997) Nature Genet. 16:303.
Citations for Recombinant Human Proprotein Convertase 1/PCSK1 Protein, CF
R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.
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Citations: Showing 1 - 2
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Ferric ions inhibit proteolytic processing of progastrin.
Authors: Bramante G, Patel O, Shulkes A, Baldwin GS
Biochem. Biophys. Res. Commun., 2010-12-30;404(4):1083-7.
Species: Human
Sample Types: Recombinant Protein
Applications: Enzyme Assay -
A proprotein convertase-inhibiting serpin with an endoplasmic reticulum targeting signal from Branchiostoma lanceolatum, a close relative of vertebrates.
Authors: Bentele C, Kruger O, Todtmann U, Oley M, Ragg H
Biochem. J., 2006-05-01;395(3):449-56.
Species: Mouse
Sample Types: Whole Cells
Applications: Bioassay
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