Recombinant Cynomolgus Monkey DPPIV/CD26 Protein, CF Summary
Product Specifications
Asn29-Pro766 with a C-terminal 6-His tag
Analysis
Product Datasheets
Carrier Free
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.
9637-SE
| 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.
|
Assay Procedure
- Assay Buffer: 25 mM Tris, pH 8.0
- Recombinant Cynomolgus Monkey DPPIV/CD26 (rcynoDPPIV) (Catalog # 9637-SE)
- Substrate: H-Gly-Pro-AMC (Bachem, Catalog # I-1225), 10 mM stock in DMSO
- F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
- Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
- Dilute rcynoDPPIV to 0.2 ng/µL in Assay Buffer.
- Dilute Substrate to 20 µM in Assay Buffer.
- Load 50 μL of 0.2 ng/µL rcynoDPPIV into a plate, and start the reaction by adding 50 μL of 20 µM Substrate. For Substrate Blank, load 50 μL of Assay Buffer and 50 μL of Substrate.
- Read at excitation and emission wavelengths of 380 nm and 460 nm (top read), respectively, in kinetic mode for 5 minutes.
- 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 7-amino, 4-Methyl Coumarin (Sigma, Catalog # A9891).
Per Well:- rcynoDPPIV: 0.010 μg
- Substrate: 10 µM
Reconstitution Calculator
Background: DPPIV/CD26
DPPIV/CD26 is an approximately 110 kDa serine exopeptidase that releases Xaa-Pro or Xaa-Ala dipeptides from the N-terminus of oligo- and polypeptides. It regulates immune and endocrine function through the cleavage of multiple chemokines, growth factors, and peptide hormones (1, 2). Mature DPPIV consists of a cytoplasmic tail, a transmembrane segment, and an extracellular domain (ECD) that contains the catalytic active site (Ser, Asp, and His charge relay system) (3). Within the ECD, cynoDPPIV/CD26 shares 96% amino acid sequence identity with human DPPIV. DPPIV is expressed as a nocovalent homodimer on the surface of epithelial cells, endothelial cells, and activated lymphocytes, and it can be released by MMP mediated shedding (4). It cleaves a range of peptide hormones including Glucagon, Glucagon-like Peptides 1 and 2, GIP, GHRH, Procalcitonin, Neuropeptide Y, and Substance P (5). It is released from adipocytes and induces insulin resistance in adipocytes and skeletal muscle (6). DPPIV also cleaves many chemokines, resulting in reduced chemotactic activity of CXCL6, 9, 10, 11, 12, and CCL5 (7-10) but unchanged angiostatic activity of CXCL9 and CXCL10 (8). Cleavage can increase (CCL5), decrease (CXCL12), or have no effect (CCL4) on chemokine blockade of HIV-1 cellular infectivity (7, 9, 11). In addition, DPPIV cleavage of CCL4 broadens chemokine receptor usage to also include CCR2b (11). DPPIV serves as a cell entry coreceptor for HIV and coronavirus (12, 13). It cleaves human GM-CSF and IL-3 and reduces their ability to promote myeloid cell development (14). It also interferes with CXCL12 induced hematopoietic cell migration, homing, and engraftment (15). DPPIV interacts in cis with Adenosine Deaminase on T cells and in trans with Caveolin-1 on antigen presenting cells (16, 17). It provides costimulatory proliferation and activation signals to both CD4+ and CD8+ T cells (17, 18).
- Klemann, C. et al. (2016) Clin. Exp. Immunol. 185:1.
- Metzemaekers, M. et al. (2016) Front Immunol. 7:483.
- Tanaka, T. et al. (1992) J. Immunol. 149:481.
- Rohrborn, D. et al. (2014) FEBS Lett. 588:3870.
- Waumans, Y. et al. (2015) Front. Immunol. 6:387.
- Lamers, D. et al. (2011) Diabetes 60:1917.
- Proost, P. et al. (1998) J. Biol. Chem. 273:7222.
- Proost, P. et al. (2001) Blood 98:3554.
- Ohtsuki, T. et al. (1998) FEBS Lett. 431:236.
- Barreira da Silva, R. et al. (2015) Nat. Immunol. 16:850.
- Guan, E. et al. (2002) J. Biol. Chem. 277:32348.
- Callebaut, C. et al. (1993) Science 262:2045.
- Raj, V.S. et al. (2013) Nature 495:251.
- Broxmeyer, H.E. et al. (2012) Nat. Med. 18:1786.
- Christopherson II, K.W. et al. (2004) Science 305:1000.
- Kameoka, J. et al. (1993) Science 261:466.
- Ohnuma, K. et al. (2007) J. Biol. Chem. 282:10117.
- Hatano, R. et al. (2013) Immunology 138:165.
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