Recombinant Human Proprotein Convertase 2/PCSK2 Protein, CF

Catalog # Availability Size / Price Qty
6018-SE-010
R&D Systems Recombinant Proteins and Enzymes
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Recombinant Human Proprotein Convertase 2/PCSK2 Protein, CF Summary

Product Specifications

Purity
>90%, by SDS-PAGE under reducing conditions and visualized by silver stain.
Endotoxin Level
<1.0 EU per 1 μg of the protein by the LAL method.
Activity
Measured by its ability to cleave the fluorogenic peptide substrate pERTKR-AMC (Catalog # ES013). The specific activity is >180 pmol/min/μg, as measured under the described conditions.
Source
Chinese Hamster Ovary cell line, CHO-derived human Proprotein Convertase 2/PCSK2 protein
Gly110-Asn638 with a C-terminal 6-His tag
Accession #
N-terminal Sequence
Analysis
Gly110
Predicted Molecular Mass
59 kDa
SDS-PAGE
66-82 kDa, reducing conditions

Product Datasheets

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6018-SE

Carrier Free

What does CF mean?

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.

What formulation is right for me?

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.

6018-SE

Formulation Supplied as a 0.2 μm filtered solution in Tris, NaCl and CaCl2.
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.
  • 6 months from date of receipt, -70 °C as supplied.
  • 3 months, -70 °C under sterile conditions after opening.

Assay Procedure

Materials
  • Assay Buffer: 50 mM NaOAc, 100 mM NaCl, 1 mM CaCl2, 0.5% (w/v) Brij-35, pH 5.0
  • Recombinant Human Proprotein Convertase 2/PCSK2 (rhPCSK2) (Catalog # 6018-SE)
  • Substrate: L-PYROGlu-Arg-Thr-Lys-Arg-AMC (Catalog # ES013), 4 mM stock in deionized water
  • F16 Black Maxisorp Plate (Nunc, Catalog # 475515
  • Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
  1. Dilute rhPCSK2 to 0.4 µg/mL in Assay Buffer.
  2. Dilute Substrate to 500 µM in Assay Buffer.
  3. Start the reaction by mixing equal volumes of 0.4 µg/mL rhPCSK2 and 500 µM Substrate together and incubate at 37 °C for 30 minutes. For a Substrate Blank, mix Assay Buffer and Substrate together and incubate at 37 °C for 30 minutes.
  4. Into a plate, load 100 µL of the reaction and Substrate Blank (in triplicate).
  5. Read at excitation and emission wavelengths of 380 nm and 460 nm (top read), respectively, in endpoint mode.
  6. 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 # A-9891).

Per Well:
  • rhPCSK2: 0.02 µg
  • Substrate: 250 µM
Reconstitution Calculator

Reconstitution Calculator

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Background: Proprotein Convertase 2/PCSK2

Proprotein Convertase 2 (PCSK2) is a subtilisin-like serine peptidase that processes proteins into biologically active products in the secretory pathway. Like other furin/kexin family members, PCSK2 cleaves its substrates primarily after sites of paired basic amino acid residues (1). Autoactivation of PCSK2 occurs in a post‑Golgi compartment of the secretory system (2). Interaction with the secretory protein 7B2 is required for proper activation of PCSK2 (3). The N-terminal domain of 7B2 stabilizes active PCSK2, and a C-terminal fragment of 7B2 can inhibit PCSK2 activity (4). rhPCSK2 was coexpressed with 7B2 to facilitate activation of the enzyme. PCSK2 is a major proteolytic processing enzyme of the regulated secretory pathway of the neuroendocrine system, where it generates a number of hormones and neuropeptides (5). Products of PCSK2 processing include enkephalins, insulin, somatostatin, dynorphin, and LHRH. 

References
  1. Zhou, A. et al. (1999) J. Biol. Chem. 274:20745.
  2. Lamango, N.S. et al. (1999) Arch. Biochem. Biophys. 362:275.
  3. Zhu, X. and I. Lindberg (1995) J. Cell Biol. 129:1641.
  4. Lamango, N. et al. (1996) Arch. Biochem. Biophys. 330:238.
  5. Rouillé, Y. et al. (1995) Front. Neuroendocrinol. 16:322.
Long Name
Proprotein Convertase Subtilisin/Kexin Type 2
Entrez Gene IDs
5126 (Human); 18549 (Mouse); 25121 (Rat)
Alternate Names
EC 3.4.21; EC 3.4.21.94; KEX2-like endoprotease 2; NEC 2; NEC2; NEC2SPC2; neuroendocrine convertase 2; PC2; PC2NEC-2; PCSK2; Prohormone convertase 2; Proprotein Convertase 2; proprotein convertase subtilisin/kexin type 2; SPC2

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Fluorogenic Peptide Substrates

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