Recombinant Human PCPE-1 Protein, CF

• Reactivity: Hu
• Applications: Enzyme Activity
• Format: Carrier-Free

Recombinant Human PCPE-1 Protein, CF Summary

• Details of Functionality: Measured by the stimulation of procollagen type I processing by BMP-1. >50% of the full-length procollagen, in the presence of Recombinant Human PCPE-1, is processed by BMP-1, as measured under the described conditions.
• Source: Mouse myeloma cell line, NS0-derived human PCPE-1 protein
  Met1-Asp449, with a C-terminal 10-His tag
• Accession #: Q15113
• N-terminal Sequence: No result, Gln26 expected
• Protein/Peptide Type: Recombinant Enzymes
• Gene: PCOLCE
• Purity: >95%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.
• Endotoxin Note: <1.0 EU per 1 μg of the protein by the LAL method.

Applications/Dilutions

• Dilutions:
  • Enzyme Activity
• Theoretical MW: 47 kDa.
  Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.
• SDS-PAGE: 51-57 kDa, reducing conditions

Packaging, Storage & Formulations

• 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.
• Buffer: Supplied as a 0.2 μm filtered solution in MES and NaCl.
• Purity: >95%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.
• Assay Procedure:
  • Dilution Buffer: 50 mM Tris, 5 mM CaCl₂, pH 7.5
  • Assay Buffer: 50 mM Tris, 5 mM CaCl₂, 150 mM NaCl, pH 7.5 (TCN)
  • Recombinant Human PCPE-1 (rhPCPE-1) (Catalog # 2627-PE)
  • Recombinant Human Pro Collagen I alpha 1 (rhCOL1A1) (Catalog # 6220-CL)
  • Recombinant Human BMP‑1/PCP (rhBMP-1) (Catalog # 1927-ZN)
  • Reducing SDS-PAGE Sample Buffer (3X or greater)
  • SDS-PAGE or Western Blot
  1. Dilute rhCOL1A1 to 75 µg/mL in Dilution Buffer.
  2. Dilute rhPCPE-1 to 15 µg/mL in Assay Buffer.
  3. In a tube, combine equal volumes (a minimum of 15 µL of each is required for this assay) of diluted rhCOL1A1 and diluted rhPCPE‑1. Prepare samples in duplicate (one replicate to be used as a control).
  4. Incubate vials at 37 °C for 30 minutes.
  5. Dilute rhBMP-1 to 1.875 µg/mL in Assay Buffer.
  6. Add one equivalent (1/2 the total volume of rhCOL1A1/rhPCPE-1 mixtures) of diluted rhBMP-1 to each reaction tube. Prevent the reaction from occurring in the control duplicates by immediately mixing with reducing gel buffer (see step 8 for volumes).
  7. Incubate reaction tubes at 37 °C for 30 minutes.
  8. After incubation, combine 40 µL of each reaction mixture and control with 20 µL of reducing SDS-PAGE gel buffer. Mix and incubate samples at 95-100 °C for 3-5 minutes.
  9. Load 30 µL per lane and analyze the cleavage by SDS-PAGE followed by protein staining and/or Western blot.
  10. Activity calculation:

  % Cleavage = [1 -	% full-length rhCOL1A1 (reaction)	] x 100%
  	% full-length rhCOL1A1 (control)

  Per Lane:
  • rhPCPE-1: 0.1 µg
  • rhCOL1A1: 0.5 µg
  • rhBMP-1: 0.0125 µg

Notes

Coomassie is a registered trademark of Imperial Chemical Industries Ltd.

This product is produced by and ships from R&D Systems, Inc., a Bio-Techne brand.

Alternate Names for Recombinant Human PCPE-1 Protein, CF

• PCOLCE
• PCPE
• PCPE1
• PCPE-1
• PCPE1Type I procollagen COOH-terminal proteinase enhancer
• procollagen C-endopeptidase enhancer 1
• procollagen C-endopeptidase enhancer
• Procollagen COOH-terminal proteinase enhancer 1
• Procollagen C-proteinase enhancer 1
• procollagen, type 1, COOH-terminal proteinase enhancer
• Type 1 procollagen C-proteinase enhancer protein

Background

Procollagen C‑endopeptidase enhancers, known as PCPEs or PCOLCEs, are secreted extracellular matrix glycoproteins that consist of two CUB domains and one NTR domain. They are known to stimulate enzymatic cleavage of procollagens I‑III by the BMP‑1/tolloid family of metalloproteases, also known as procollagen C‑proteinases (1). PCPE‑1 is expressed primarily by interstitial connective tissues such as tendons, calvaria, and skin (2). Although BMP‑1/tolloid proteinases are involved in processing of multiple extracellular proteins, the enhancer activity of PCPE‑1 is specific to procollagens since it has no effect on BMP‑1/tolloid cleavage of other substrates (3). It is thought that PCPE‑1 enhances cleavage of procollagens by binding to the substrate and inducing a conformation change in the substrate (3), although interaction between PCPE‑1 and full‑length BMP‑1/tolloid proteinases has also been reported (4).

1. Steiglitz, B.M. et al. (2002) J. Biol. Chem. 277:49820.
2. Kessler, E. et al. (1990) Biochem. Biophys. Res. Commun. 173:81.
3. Moali, C. et al. (2005) J. Biol. Chem. 280:24188.
4. Ge, G. et al. (2006) J. Biol. Chem. 281:10786.

Bioinformatics

• Gene Symbol: PCOLCE
• Uniprot:
  • Q15113()