Recombinant Human IMP Dehydrogenase 2/IMPDH2 Protein, CF

Catalog # Availability Size / Price Qty
8349-DH-050
Recombinant Human IMP Dehydrogenase 2/IMPDH2 Protein, CF Enzyme Activity
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Recombinant Human IMP Dehydrogenase 2/IMPDH2 Protein, CF Summary

Purity
>90%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.
Endotoxin Level
<0.10 EU per 1 μg of the protein by the LAL method.
Activity
Measured by its ability to convert the substrate inosine-5'-phosphate (IMP) to xanthosine-5'-phosphate (XMP). The specific activity is >30 pmol/min/μg, as measured under the described conditions.
Source
Spodoptera frugiperda, Sf 21 (baculovirus)-derived human IMP Dehydrogenase 2/IMPDH2 protein
Met1-Phe514, with a C-terminal 10-His tag
Accession #
N-terminal Sequence
Analysis
Met1 predicted
Predicted Molecular Mass
57 kDa
SDS-PAGE
52-61 kDa, reducing conditions

Product Datasheets

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.

8349-DH

Formulation Supplied as a 0.2 μm filtered solution in Tris, NaCl, DTT and Glycerol.
Shipping The product is shipped with dry ice or equivalent. 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, 300 mM NaCl, 1 mM EDTA, 1 mM DTT, pH 8.0
  • Recombinant Human Inosine 5'-Monophosphate Dehydrogenase 2/IMPDH2 (rhIMPDH2) (Catalog # 8349-DH)
  • Nicotinamide adenine dinucleotide sodium salt ( beta -NAD) (Sigma, Catalog # N6522), 100 mM stock in deionized water
  • Inosine 5'-monophosphate (IMP) (Sigma, Catalog # I4625), 100 mM stock in deionized water.
  • UV plate (Costar, Catalog # 3635)
  • Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
  1. Dilute rhIMPDH2 to 20 μg/mL in Assay Buffer.
  2. Create Substrate Mixture containing 500 μM IMP and 1 mM beta -NAD in Assay Buffer.
  3. Load 50 μL of 20 μg/mL rhIMPDH-2 into a plate, and start the reaction by adding 50 μL of Substrate Mixture. For Substrate Blanks, load 50 μL of Assay Buffer and 50 μL of Substrate Mixture.
  4. Read plate at a wavelength of 339 nm (bottom read) in kinetic mode for 5 minutes.
  5. Calculate specific activity:

     Specific Activity (pmol/min/µg) =

Adjusted Vmax* (OD/min) x well volume (L) x 1012 pmol/mol
ext. coeff** (M-1cm-1) x path corr.*** (cm) x amount of enzyme (µg)

 

*Adjusted for Substrate Blank.
**Using the extinction coefficient 6220 M-1cm-1.
***Using the path correction 0.320 cm.
Note: the output of many spectrophotometers is in mOD.

Per Well:
  • rhIMPDH-2: 1.0 μg
  • beta -NAD: 500 μM
  • IMP: 250 μM

Data Images

Enzyme Activity View Larger

Recombinant Human IMPDH2 (Catalog # 8349-DH) is measured by its ability to convert the substrate inosine-5'-phosphate (IMP) to xanthosine-5'-phosphate (XMP). The activity (orange) is approximately 3.5-fold greater than the competitor's IMPDH2 (green).

SDS-PAGE View Larger

1 μg/lane of Recombinant Human Inosine 5'-Monophosphate Dehydrogenase 2/IMPDH2 was resolved with SDS-PAGE under reducing (R) and non-reducing (NR) conditions and visualized by silverstaining, showing a band at 57 kDa.

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: IMP Dehydrogenase 2/IMPDH2

IMPDH2 (inosine monophosphate dehydrogenase) is one of two cytosolic and nuclear enzyme isoforms that play a central role in guanine nucleotide metabolism. The isoforms are 84% identical but distinctly regulated. While IMPDH1 is generally constitutively expressed, IMPDH2 is inducible during proliferation and transformation (1, 2). Both isoforms form a homotetramer of approximately 55 kDa monomers containing a catalytic barrel domain and a subdomain with two cystathione beta-synthase domains which mediate RNA and DNA binding (1, 3). Both enzymes catalyze the NAD-dependent conversion of inosine monophosphate (IMP) to hypoxanthine monophosphate (XMP) which is a precursor for the synthesis of GMP, guanosine, and guanine. These compounds are critical for DNA synthesis and cell proliferation (4, 5) which explains the importance of IMPDH in cancer and viral infection (6-9). Although IMPDH1 and IMPDH2 are known to be inhibited by the immunosuppressant drug mycophenolic acid (MPA), much research has targeted discovery of additional and selective inhibitors for IMPDH isoforms given they are targets for several major therapeutic areas (1, 9-13). Human IMPDH2 shares 99% amino acid sequence identity with mouse IMPDH2.

References
  1. Hedstrom, L. (2009) Chem. Rev. 109:2903.
  2. Thomas, E.C. et al. (2012) PLoS One 12:e51096.
  3. McLean, J.E. et al. (2004) Biochem. J. 379:243.
  4. Lane, A.N. and T.W. Fan (2015) Nucleic Acids Res. 43:2466.
  5. Jackson, R.C. et al. (1975) Nature 256:331.
  6. Zho, J. et al. (2015) Med. Oncol. 32:373.
  7. Xu, Y. et al. (2017) Sci. Rep. 7:745.
  8. Nair, V. (2007) Antivir. Chem. Chemother. 18:245.
  9. Trapero, A. et al. (2018) J. Med. Chem. 61:2806.
  10. Barnes, B.J. et al. (2001) Int. J. Cancer 94:275.
  11. Cholewinksi, G. et al. (2015) J. Enzyme Inhib. Med. Chem. 30:550.
  12. Liao, L.X. et al. (2017) Proc. Natl. Acad. Sci. USA 114:E5986.
  13. Cuny, G.D. et al. (2017) Expert Opin. Ther. Pat. 27:677.
Long Name
Inosine 5'-Monophosphate Dehydrogenase 2
Entrez Gene IDs
3615 (Human); 23918 (Mouse); 301005 (Rat)
Alternate Names
EC 1.1.1.205; IMDH2; IMP (inosine 5'-monophosphate) dehydrogenase 2; IMP (inosine monophosphate) dehydrogenase 2; IMP Dehydrogenase 2; IMP Oxireductase 2; IMPD 2; IMPD2; IMPD2IMP oxireductase 2; IMPDH 2; IMPDH2; IMPDH-II; inosine 5' phosphate dehydrogenase 2; inosine monophosphate dehydrogenase type II; inosine-5'-monophosphate dehydrogenase 2

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