Recombinant Human Glutaminase His-tag Protein, CF

R&D Systems | Catalog # 10115-GL

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Key Product Details

  • R&D Systems E. coli-derived Recombinant Human Glutaminase His-tag Protein (10115-GL)
  • Quality control testing to verify active proteins with lot specific assays by in-house scientists
  • All R&D Systems proteins are covered with a 100% guarantee

Source

E. coli

Accession Number

O94925-1

Applications

Enzyme Activity
View all Glutaminase Proteins and Enzymes »

Product Specifications

Source

E. coli-derived human Glutaminase protein
Ser17-Leu669
with N-terminal Met and 6-His tag

Purity

>80%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.

N-terminal Sequence Analysis

Met

Predicted Molecular Mass

73 kDa

SDS-PAGE

59-74 kDa, under reducing conditions

Activity

Measured by its ability to hydrolyze glutamine to glutamate.
The specific activity is >12000 pmol/min/μg, as measured under the described conditions.

Scientific Data Images for Recombinant Human Glutaminase His-tag Protein, CF

Recombinant Human Glutaminase His-tag Protein Enzyme Activity

Recombinant Human Glutaminase His-tag Protein Enzyme Activity

Recombinant Human Glutaminase His-tag (Catalog # 10115-GL) is measured by its ability to hydrolyze glutamine to glutamate. The activity (orange) is higher than the competitor's Glutaminase (green).
Recombinant Human Glutaminase His-tag Protein SDS-PAGE

Recombinant Human Glutaminase His-tag Protein SDS-PAGE

1 μg/lane of Recombinant Human Glutaminase His-tag (Catalog # 10115-GL) and 1 μg/lane of competitor Human Glutaminase was resolved with 4-20% SDS-PAGE under reducing (R) and non-reducing (NR) conditions and visualized by silver staining.

Formulation, Preparation, and Storage

10115-GL
Formulation Supplied as a 0.2 μm filtered solution in Tris, NaCl, TCEP, Glycerol 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.
  • 6 months from date of receipt, -70 °C as supplied.
  • 3 months, -70 °C under sterile conditions after opening.

Background: Glutaminase

Glutaminase (GLS) catalyzes the synthesis of glutamate from glutamine. There are two mammalian isozymes of glutaminase: kidney-type encoded by the GLS gene, and liver-type encoded by the GLS2 gene. The two isozymes are known to have different localization, functionality, and immunological and molecular characteristics (1, 2). The GLS gene expresses two splice variants with a conserved catalytic domain (3): kidney-type glutaminase (KGA) and a C-terminal truncated variant known as glutaminase C. Both splice variants form active tetramers and are activated by phosphate (4). The kidney glutaminase isoform, GLS, catalyzes the first reaction in the primary pathway for renal catabolism of glutamine to glutamate. It contains a mitochondrial signaling peptide, a nuclear receptor box, a catalytic glutaminase domain, three ankyrin repeats and a KEN box (5).  GLS promotes respiration and ATP generation in the mitochondria by providing an alternative input to the citric acid cycle. GLS is upregulated in response to oncogenes (5-7) in several cancers (8, 9). Cancer cells with altered metabolism have been shown to depend on GLS activity and upregulated glutamine consumption (10) for growth in acute myeloid leukemina (11), breast cancer (12), colorectal cancer (13), kidney cancer (14), lung cancer (15), melanoma (16) and pancreatic cancer (17). Based on its role in cancer, GLS is a pharmaceutical target for cancer therapy (10, 12, 13, 18).

References

  1. Kovacevic, Z. et al. (1983) Physiol. Rev. 63:547.
  2. Marquez, J. et al. (2006) Neurochem. Int. 48:465.
  3. Ramachandran, S. et al. (2016) Oncotarget. 7:57943.
  4. Huang, Q. et al. (2018) J. Biol. Chem. 293:3535.
  5. Thangavelu, K. et al. (2012) Proc. Natl. Acad. Sci. 109:7705.
  6. Wise, D. et al. (2008) Proc. Natl. Adac. Sci. 105:18782.
  7. Gao, P. et al. (2009) Nature. 458:762.
  8. Cline, M. S. et al. (2013) Sci. Rep. 3:2652.
  9. Cancer Genome Atlas Research Network, et al. (2013) Nat. Genet. 45:1113.
  10. Katt, W. P. et al. (2017) Future Med. Chem. 9:223.
  11. Willems, L. et al. (2013) Blood 122:3521.
  12. Gross, M. I. et al. (2014) Mol. Cancer Ther. 13:890.
  13. Huang, F. et al. (2014) Int. J. Clin. Exp. Pathol. 7:1093.
  14. Shroff, E. H. et al. (2015) Proc. Natl. Acad. Sci. 112:6539.
  15. van den Heuvel, A. P. et al. (2012) Cancer Biol. Ther. 13:1185.
  16. Hernandez-Davies, J. E. et al. (2015) J. Trans. Med. 13:1.
  17. Son, J. et al. (2013) Nature. 496:101.
  18. Zimmermann, S.C. et al. (2018) J. Med. Chem. [Epub ahead of print].

Long Name

Glutaminase Kidney Isoform, Mitochondrial

Alternate Names

GLS, K-Glutaminase, L-Glutamine Amidohydrolase

Entrez Gene IDs

2744 (Human); 14660 (Mouse); 24398 (Rat)

Gene Symbol

GLS

UniProt

O94925-1

Additional Glutaminase Products

Product Documents for Recombinant Human Glutaminase His-tag Protein, CF

Certificate of Analysis

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Product Specific Notices for Recombinant Human Glutaminase His-tag Protein, CF

For research use only

Citations for Recombinant Human Glutaminase His-tag Protein, CF

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Protocols

View specific protocols for Recombinant Human Glutaminase His-tag Protein, CF (10115-GL):

Materials
  • Assay Buffer: 100 mM Tris, 150 mM K2HPO4, pH 8.6
  • Recombinant Human Glutaminase Kidney Isoform His-tag (rhGLS) (Catalog # 10115-GL)
  • Glutamate dehydrogenase (GIDH) (Sigma, Catalog # G7882), 400 U/mL stock in 10 mM HEPES, pH 7.0
  • Nicotinamide adenine dinucleotide ( beta -NAD) (Sigma, Catalog # N6522), 100 mM stock in deionized water
  • L-Glutamine (Sigma, Catalog # G8540), 200 mM stock in deionized water
  • 96-well Clear Plate (Catalog # DY990)
  • Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
  1. Dilute rhGLS to 0.5 µg/mL in Assay Buffer.
  2. Prepare Substrate Mixture containing 50 mM L-Glutamine, 60 U/mL GIDH and 4 mM beta -NAD in Assay Buffer.
  3. Load into a plate 50 µL of rhGLS, and start the reaction by adding 50 µL of Substrate Mixture.  Include a Substrate Blank containing 50 µL Assay Buffer and 50 µL of Substrate Mixture.
  4. Read plate at 340 nm (absorbance) 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 0.32 cm.
Note: the output of many spectrophotometers is in mOD. Per Well:

  • rhGLS: 0.025 µg
  • L-Glutamine: 25 mM
  • GIDH: 3 U
  • beta -NAD: 2 mM

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