Recombinant Mouse PLA2G2A Protein, CF

Catalog # Availability Size / Price Qty
4925-PL-010
R&D Systems Recombinant Proteins and Enzymes
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Recombinant Mouse PLA2G2A Protein, CF Summary

Product Specifications

Purity
>95%, 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 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 #
N-terminal Sequence
Analysis
Asn22
Predicted Molecular Mass
15 kDa
SDS-PAGE
17 kDa, reducing conditions

Product Datasheets

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4925-PL

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.

4925-PL

Formulation Supplied as a 0.2 μm filtered solution in Sodium Acetate 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.

Assay Procedure

Materials
  • 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
Reconstitution Calculator

Reconstitution Calculator

The reconstitution calculator allows you to quickly calculate the volume of a reagent to reconstitute your vial. Simply enter the mass of reagent and the target concentration and the calculator will determine the rest.

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Background: PLA2G2A

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).

References
  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.
Long Name
Phospholipase A2 Group IIA
Entrez Gene IDs
5320 (Human); 18780 (Mouse); 29692 (Rat)
Alternate Names
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

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