Recombinant Mouse Pappalysin-1/PAPP-A His-tag Protein, CF Summary
Recombinant Mouse Pappalysin-1/PAPPA will cleave >50% of
Human IGFBP‑5 (Catalog # 875-B5), as measured under the described conditions.
with a C-terminal 6-His tag
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.
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.
|Formulation||Supplied as a 0.2 μm filtered solution in Tris, CaCl2, NaCl and CHAPS.|
|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.
- Assay Buffer: 50 mM Tris, 10 mM CaCl2, 150 mM NaCl (TCN)
- Recombinant Mouse Pappalysin-1/PAPPA (rmPappalysin-1) (Catalog # 10174-ZN)
- Recombinant Human IGFBP‑5 (Catalog # 875-B5)
- Reducing SDS-PAGE Gel Buffer
- SDS-PAGE or Western Blot
- Dilute rmPappalysin-1 to 10 µg/mL in Assay Buffer.
- Dilute rhIGFBP-5 to 200 µg/mL in Assay Buffer.
- Combine equal volumes of 10 µg/mL of rmPappalysin-1 and 200 µg/mL rhIGFBP-5. Include two controls containing Assay Buffer in place of rmPappalysin-1.
- Incubate reaction vials and one control at 37 C for 1 hour. Keep the other control at -20 °C during the incubation period.
- Combine equal volumes of rmPappalysin-1/rhIGFBP-5 reaction mixture and reducing SDS-PAGE gel buffer.
- Analyze the cleavage by SDS-PAGE (load 20 µL/Lane) followed by protein staining and/or Western blot.
- rmPappalysin-1/PAPPA: 0.050 µg
- rhIGFBP-5: 1 µg
Pappalysins belong to a fifth family of metzincins that consists of ADAMs/ADAMTSs, MMPs, astacins and serralysins (1). Pappalysin-1 (PAPP-A) is a secreted, disulfide-linked 400 kDa homodimer with an N-terminal catalytic domain and five C-terminal Sushi domains (2). PAPP-A cleaves Insulin-like Growth Factor-Binding Protein-4 and -5 (IGFBP-4 and -5) at a single site, resulting in the release of bioactive IGF (3). PAPP-A also contains three Lin12-Notch repeats (LNR) that bind calcium and are required for cleavage of IGFBP-4, but not IGFBP-5 (2). PAPP-A is thought to be the only physiological IGFBP-4 proteinase supported by the observation that no IGFBP-4 cleavage occurred in embryonic fibroblasts derived from PAPP-A knockout mice (4). As a regulator of IGF bioavailability, and consequently IGF signaling, PAPP-A is an attractive therapeutic target (5-8). PAPP-A knockout mice have an extended lifespan (9) and use of specific neutralizing antibodies causes inhibition of atherosclerotic plaque progression (10). PAPP-A levels are elevated in cancer (11-14) and PAPP-A has been identified as a migration/invasion-promoting gene (5,6, 12). PAPP-A is also important during pregnancy where PAPP-A levels are increased in the plasma by a factor of about 150. Low maternal serum levels of PAPP-A is used as an indication of Down syndrome or other fetal aneuploidies in the first trimester of pregnancy (15).
- Boldt, H. B. et al. (2001) Biochem. J. 358:359.
- Boldt, H. B. et al. (2004) J. Biol. Chem. 279:38525.
- Laursen, L. S. et al. (2001) FEBS Lett. 504:36.
- Conover, C. A. et al. (2004) Development 131:1187.
- Huang, J. et al. (2013) Oncotarget. 4:1172.
- Prithviraj, P. et al. (2015) Oncotarget 6: 15953.
- Yu, X. E. et al. (2018) Atherosclerosis. 278:250.
- Guo, Y. et al. (2018) Am. J. Cancer Res. 8:955.
- Conover, C. A. et al. (2010) J. Gerontol. A. Biol. Sci. Med. Sci. 65:590.
- Conover, C. A. et al. (2016) J Cardiovasc. Transl. Res. 9:77.
- Bulut, I. et al. (2009) Am. J. Med. Sci. 337: 241.
- Boldt, H. B. and Conover C. A. (2011) Endocrinology 152:1470.
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