Recombinant Human Pepsinogen A5/PGA5 Protein, CF

R&D Systems | Catalog # 8457-AS

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Key Product Details

  • R&D Systems HEK293-derived Recombinant Human Pepsinogen A5/PGA5 Protein (8457-AS)
  • Quality control testing to verify active proteins with lot specific assays by in-house scientists
  • All R&D Systems proteins are covered with a 100% guarantee

Source

HEK293

Accession Number

P0DJD9

Applications

Enzyme Activity
View all Pepsinogen A5/PGA5 Proteins and Enzymes »

Product Specifications

Source

Human embryonic kidney cell, HEK293-derived human Pepsinogen A5/PGA5 protein
Ile16-Ala388, with C-terminal 6-His tag
Accession # P0DJD9

Purity

>95%, by SDS-PAGE under reducing conditions and visualized by Colloidal Coomassie® Blue stain at 5 μg per lane.

Endotoxin Level

<1.0 EU per 1 μg of the protein by the LAL method.

N-terminal Sequence Analysis

Ile16 & Val63

Predicted Molecular Mass

41 kDa

SDS-PAGE

39-51 kDa, reducing conditions

Activity

Measured by its ability to cleave the fluorogenic peptide substrate, Mca-RPKPVE-Nval-WRK(Dnp)-NH2 (Catalog # ES002).
The specific activity is >15,500 pmol/min/μg, as measured under the described conditions.

Formulation, Preparation, and Storage

8457-AS
Formulation Supplied as a 0.2 μm filtered solution in Tris 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.
  • 6 months from date of receipt, -20 to -70 °C as supplied.
  • 3 months, -20 to -70 °C under sterile conditions after opening.

Background: Pepsinogen A5/PGA5

Pepsins are aspartic proteases that are synthesized in the gastric mucosa and secreted into the stomach. They are released as zymogens called pepsinogens which are then converted to active pepsins by the acidic pH of gastric juices (1-3). PGA3, PGA4, and PGA5 are human Pepsinogen A isozymogens that differ in sequence by 2­4 amino acid (aa) residues (4, 5). This recombinant human Pepsinogen A corresponds to PGA5. Human Pepsinogen A isozymogens share approximately 56% aa sequence identity with mouse and rat Pepsinogen A isozymogens. Pepsins have optimal activity under conditions of acidic pH and are inhibited by pepstatin (6, 7). Pepsin A has broad substrate specificity, but preferentially cleaves peptide bonds involving aromatic and aliphatic amino acids.

References

  1. Athauda, S.B. et al. (1989) J. Biochem. 106:920.
  2. Kageyama, T. et al. (1989) J. Biochem. 105:15.
  3. Kageyama, T. (2002) Cell. Mol. Life Sci. 59:288.
  4. Zwiers, A. et al. (1994) Clin. Nephrol. 41:153.
  5. Nakai, H. et al. (1986) Cytogenet. Cell Genet. 43:215.
  6. Marciniszyn, J., Jr. et al. (1976) J. Biol. Chem. 251:7088.
  7. Kageyama, T. and K. Takahashi (1980) J. Biochem. 88:571.

Alternate Names

PGA5

Entrez Gene IDs

5222 (Human)

Gene Symbol

PGA5

UniProt

P0DJD9

Additional Pepsinogen A5/PGA5 Products

Product Documents for Recombinant Human Pepsinogen A5/PGA5 Protein, CF

Certificate of Analysis

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Note: Certificate of Analysis not available for kit components.

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Product Specific Notices for Recombinant Human Pepsinogen A5/PGA5 Protein, CF

For research use only

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Protocols

View specific protocols for Recombinant Human Pepsinogen A5/PGA5 Protein, CF (8457-AS):

Materials
  • Activation Buffer: 50 mM Sodium Citrate, pH 2.5
  • Assay Buffer: 50 mM Sodium Citrate, 500 mM NaCl, 0.02% Triton® X-100, pH 4.0
  • Recombinant Human Pepsinogen A5/PGA5 (rhPGA-5) (Catalog # 8457-AS)
  • Substrate MCA-Arg-Pro-Lys-Pro-Val-Glu-NVAL-Trp-Arg-Lys(DNP)-NH2 (Catalog # ES002), 2 mM stock in DMSO
  • F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
  • Fluorescent Plate Reader (Model: Gemini EM by Molecular Devices) or equivalent
  1. Dilute rhPGA-5 to 100 µg/mL in Activation Buffer.
  2. Incubate for 15 minutes at room temperature.
  3.  Dilute activated rhPGA-5 to 0.02 µg/mL in Assay Buffer.
  4. Dilute Substrate to 20 µM in Assay Buffer.
  5. Load 50 µL of 0.02 µg/mL of rhPGA-5 into a plate, and start the reaction by adding 50 µL of 20 µM Substrate. Include a Substrate Blank containing 50 µL of Assay Buffer and 50 µL of 20 µM Substrate.
  6. Read at excitation and emission wavelengths of 320 nm and 405 nm (top read), respectively, in kinetic mode for 5 minutes.
  7. 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). Per Well:

  • rhPGA-5: 0.001 µg
  • Substrate: 10 µM

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