Recombinant Mouse PLA2G2A Protein, CF

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

Recombinant Mouse PLA2G2A Protein, CF Summary

• Details of Functionality: Measured by its ability to hydrolyze 1-Hexadecanoyl-2-(1-pyrene-decanoyl)-sn-glycero-3-phosphocholine. The specific activity is >50 pmol/min/µg, as measured under the described conditions.
• Source: Spodoptera frugiperda, Sf 21 (baculovirus)-derived mouse PLA2G2A protein
  Asn22-Cys146, with a C-terminal 10-His tag
• Accession #: NP_001076000
• N-terminal Sequence: Asn22
• Protein/Peptide Type: Recombinant Enzymes
• Gene: Pla2g2a
• Purity: >95%, by SDS-PAGE under reducing conditions and visualized by silver stain.
• Endotoxin Note: <1.0 EU per 1 μg of the protein by the LAL method.

Applications/Dilutions

• Dilutions:
  • Enzyme Activity
• Theoretical MW: 15 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: 17 kDa, reducing conditions

Packaging, Storage & Formulations

• 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.
• Buffer: Supplied as a 0.2 μm filtered solution in Sodium Acetate and NaCl.
• Purity: >95%, by SDS-PAGE under reducing conditions and visualized by silver stain.
• Assay Procedure:
  • Assay Buffer: 50 mM Tris, 500 mM NaCl, 1 mg/mL BSA, pH 8.5.
  • Substrate Buffer: 50 mM Tris, 500 mM NaCl, 20 mM CaCl₂, 1 mg/mL BSA, pH 8.5.
  • Recombinant Mouse Phospholipase A2 Group IIA/PLA2G2A (rmPLA2G2A) (Catalog # 4925-PL)
  • Substrate: 1-hexadecanoyl-2-(1-pyrene-decanoyl)-sn-glycero-3-phosphocholine (Invitrogen, Catalog # H361), Dilute to 2 mM in DMSO, then dilute to a final stock concentration of 400 µM in ethanol
  • F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
  • Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
  1. Thaw Substrate at 37 °C for five minutes. Mix well.
  2. Dilute rmPLA2G2A to 2 ng/µL in Assay Buffer.
  3. Dilute substrate PGPC to 20 µM in Substrate Buffer.
  4. Load in plate 50 µL of 2 ng/µL rmPLA2G2A, and start the reaction by adding 50 µL of 20 µM Substrate to each well. Include a Substrate Blank containing 50 µL Assay Buffer and 50 µL Substrate.
  5. Read at excitation and emission wavelengths of 345 nm and 395 nm (top read), respectively, in kinetic mode for 5 minutes.
  6. 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 1-pyrenedecanoic acid (Invitrogen, Catalog # P31).

  Per Well:
  • rmPLA2G2A: 0.1 µg
  • Substrate: 10 µM

Notes

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

Alternate Names for Recombinant Mouse PLA2G2A Protein, CF

• EC 3.1.1.4
• GIIC sPLA2
• Group IIA phospholipase A2
• Mom1
• Non-pancreatic secretory phospholipase A2
• NPS-PLA2
• Phosphatidylcholine 2-acylhydrolase 2A
• phospholipase A2, group IIA (platelets, synovial fluid)
• phospholipase A2, membrane associated
• PLA2B
• PLA2BPLA2
• PLA2G2A
• PLA2L
• PLA2LPLA2S
• PLAS1
• RASF-A
• sPLA2

Background

Secretory Phospholipase A₂ is an enzyme that hydrolyses the sn-2 ester bond of phospholipids, generating non-esterified free fatty acids and lysophospholipids (1‑3). PLA2G2A is a calcium-dependent phospholipase expressed in many cell types participating in inflammation-associated cellular responses, including platelets, neutrophils, and mast cells. It may function as an enzymatic component of the host defense mechanism. For example, human tears contain a high concentration of PLA2G2A, a principal bactericidal factor against Gram-positive bacteria in this fluid. It may play a role in cell proliferation through binding a receptor on the cell membrane. PLA2G2A has been shown to have pro-atherogenic properties both in the circulation and within the arterial wall (4). It is an acute phase protein expressed in response to a variety of pro-inflammatory cytokines. Circulating levels of sPLA2G2A are higher in coronary artery disease (CAD) patients and are associated with increased risk of future CAD (5).

1. Webb, N. R. (2005) Cur. Opin. Lipid. 16:341.
2. Triggiani, M. et al. (2005) J. Allergy Clin. Immunol. 116:1000.
3. Murakami, M. and Kudo, I. (2004) Biol. Pharm. Bull. 27:1158.
4. de Beer, F. C. and Webb, N. R. (2006) Arterioscler. Thromb. Vasc. Biol. 26:1421.
5. Wootton, P. T. E. et al. (2006) Human Mol. Genet. 15:355.

Bioinformatics

• Gene Symbol: Pla2g2a
• Uniprot:
  • NP_001076000()