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Recombinant Human FGFR1 Kinase Domain Protein, CF

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Summary
Reactivity HuSpecies Glossary
Applications Enzyme Activity
Format
Carrier-Free

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Recombinant Human FGFR1 Kinase Domain Protein, CF Summary

Details of Functionality
Measured by its ability to hydrolyze the 5’-phosphate groups from the substrate adenosine-5’-triphosphate (ATP). The orthophosphate product is measured by a Malachite Green Phosphate Detection Kit (Catalog # DY996). The specific activity is >12 pmol/min/μg, as measured under the described conditions.
Source
Spodoptera frugiperda, Sf 21 (baculovirus)-derived human FGF R1 protein
Met399-Arg822 with an N-terminal Met and 6-His tag
Accession #
N-terminal Sequence
No results obtained: Met predicted. Protein identity confirmed by MS analysis of tryptic fragments.
Protein/Peptide Type
Recombinant Proteins
Gene
FGFR1
Purity
>90%, by SDS-PAGE under reducing conditions and visualized by Colloidal Coomassie® Blue stain at 5 μg per lane.
Endotoxin Note
<1.0 EU per 1 μg of the protein by the LAL method.

Applications/Dilutions

Dilutions
  • Enzyme Activity
Theoretical MW
49 kDa.
Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.
SDS-PAGE
45-58 kDa, reducing conditions

Packaging, Storage & Formulations

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.
Buffer
Supplied as a 0.2 μm filtered solution in Tris, NaCl and TCEP.
Purity
>90%, by SDS-PAGE under reducing conditions and visualized by Colloidal Coomassie® Blue stain at 5 μg per lane.
Assay Procedure
  • Universal Kinase Activity Kit (Catalog # EA004)
  • 10X Assay Buffer (supplied in kit): 250 mM HEPES, 1.5 M NaCl, 100 mM MgCl2, 100 mM CaCl2, pH 7.0
  • Recombinant human FGF R1 (rhFGF R1) (Catalog # 8657-FR)
  • ATP (supplied in kit): 10 mM
  • 96-well Clear Plate (Catalog # DY990)
  • Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
  1. Prepare 1X Assay Buffer by diluting 10X stock 10 fold with deionized water.
  2. Dilute 1 mM Phosphate Standard provided by the Universal Kinase Activity Kit by adding 40 µL of the 1 mM Phosphate Standard to 360 µL of 1X Assay Buffer for a 100 µM stock. This is the first point of the standard curve.
  3. Complete the standard curve by performing six one-half serial dilutions of the 100 µM Phosphate stock using 1X Assay Buffer. The standard curve has a range of 0.078 to 5 nmol per well.
  4. Dilute ATP (supplied in kit) to 0.4 mM in 1X Assay Buffer.
  5. Dilute rhFGF R1 to 66.67 ng/µL in 1X Assay Buffer.
  6. Load 50 µL of each dilution of the standard curve into a plate. Include a curve blank containing 50 μL of Assay Buffer.
  7. Load 15 µL of 66.67 ng/µL rhFGF R1 into empty wells of the same plate as the curve. Include a Control containing 15 μL of 1X Assay Buffer.
  8. Add 25 µL of 0.4 mM ATP to all wells, excluding the standard curve. 
  9. Seal plate and incubate at 37 °C for 1 hour.
  10. Dilute Coupling Phosphatase 4 to 10 ng/µL in Assay Buffer.
  11. Add 10 µL of 10 ng/µL Coupling Phosphatase 4 to all wells, excluding the standard curve.
  12. Seal and incubate plate at room temperature for 5 minutes.
  13. Add 30 µL of the Malachite Green Reagent A to all wells. Mix briefly.
  14. Add 100 µL of deionized water to all wells. Mix briefly.
  15. Add 30 µL of the Malachite Green Reagent B to all wells. Mix and incubate sealed plate for 20 minutes at room temperature.
  16. Read plate at 620 nm (absorbance) in endpoint mode.
  17. Calculate specific activity:

     Specific Activity (pmol/min/µg) =

 Phosphate released* (nmol) x (1000 pmol/nmol)
Incubation time (min) x amount of enzyme (µg)

     *Derived from the phosphate standard curve using linear or 4-parameter fitting and adjusted for Control
     **Decoupled reaction (use 65 minutes for incubation time); no coupling rate constant

Per Reaction:
  • rhFGF R1:  1.0 µg
  • Coupling Phosphatase 4:  0.1 µg
  • ATP:  0.2 mM

Notes

This product is produced by and ships from R&D Systems, Inc., a Bio-Techne brand.

Alternate Names for Recombinant Human FGFR1 Kinase Domain Protein, CF

  • basic fibroblast growth factor receptor 1
  • BFGFR
  • bFGF-R-1
  • CD331 antigen
  • CD331
  • CEK
  • EC 2.7.10
  • EC 2.7.10.1
  • FGF R1
  • FGFBR
  • FGFR1
  • FGFR-1
  • fibroblast growth factor receptor 1
  • FLGH3
  • FLJ99988
  • Flt-2
  • FLT2H4
  • Fms-like tyrosine kinase 2
  • fms-related tyrosine kinase 2
  • H2
  • H5
  • HBGFR
  • heparin-binding growth factor receptor
  • hydroxyaryl-protein kinase
  • KAL 2
  • KAL2
  • N-SAM
  • OGD
  • Proto-oncogene c-Fgr
  • soluble FGFR1 variant 1
  • soluble FGFR1 variant 2

Background

Fibroblast growth factor receptors (FGF Rs) are single-pass transmembrane proteins with extracellular fibroblast growth factor (FGF) binding domains and cytoplasmic tyrosine-protein kinase domains (1). FGF Rs play essential roles in the regulation of embryonic development, cell proliferation, differentiation and migration (2). Upon FGF and heparan sulfate binding, the FGF Rs form homodimers and the kinase domains are activated (3, 4). The activated kinases then phosphorylate downstream effector molecules such as PLC-g1, FRS2, Gab1, and SHB, leading to the activation of several signaling cascades (5). For example, activation of PLC-g1 leads to the production of the cellular signaling molecules diacylglycerol and inositol 1,4,5-trisphosphate (6). Phosphorylation of FRS2 triggers recruitment of GRB2, Gab1, PI 3-Kinase p85, and SOS1, activation of Ras, ERK2, and ERK1, and activation of the MAP kinase and AKT1 signaling pathways (7). In this construct, the kinase domain of human FGF R1 is expressed as a constitutively active enzyme (8) and the activity was assayed using a phosphatase-coupled method (9).
  1. Johnson, D.E. et al. (1990) Mol. Cell. Biol. 10:4728.
  2. Turner, N and Grose, R. (2010) Nat. Rev. Cancer 10:116.
  3. Pantoliano, M.W. et al (1994) Biochemistry 33:10229.
  4. Ornitz, D.M. et al. (996) J. Biol. Chem. 271:15292.
  5. Eswarakumar, V.P. et al. (2005) Cytokine Growth Factor Rev. 16:139.
  6. Peters, K.G. et al. (1992) Nature 358:678.
  7. Mohammadi, M. et al. (1996) Mol. Cell. Biol. 16:977.
  8. Mohammadi, M et al (1997) Science 276:955.
  9. Wu, Z.L. (2011) PloS ONE 6:e23172.

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FAQs for FGFR1 (8657-FR). (Showing 1 - 2 of 2 FAQ).

  1. I would like to inquire whether you have samples of the following antibodies: NBP1-61338, NB600-1287, NBP1-19481, NBP1-19864; The datasheets for these antibodies show different molecular weights of the detected proteins so its no altogether clear to me which one detects the real protein. To sort this out, we would be using FGFR1-deficient cells and would very much appreciate if you could supply us with small aliquots of these antibodies.
    • FGFR1 has multiple isoforms and is subject to multiple PTM's. The molecular weight can vary greatly depending on experimental conditions and samples used. We fully guarantee all of our products for the listed applications and species. If you cannot get a product to work in an application or species stated on our datasheet, our technical service team will troubleshoot with you to get it to work. If a product still does not work after troubleshooting, you can receive a free of charge replacement product or a full refund. Due to our 100% guarantee, we do not offer free of charge samples of our products. If a smaller sample size is available, it will be listed on our product page.
  2. I am working in Multiple sclerosis group and mainly working with mice model and cell culture study. I need FGFR1 antibody against Mice for FACS analysis. If it is available can I get the sample to check with my cells? And any Oligodendrocyte markers against mice for FACS too.
    • We do have an antibody that has been tested in FLOW, NB600-1287, however it has not been tested in mouse. If you would like to test this antibody in mouse samples we have an Innovators Reward Program where we reward you for trying our antibody in species and applications that have not been previously tested.

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Bioinformatics

Gene Symbol FGFR1
Uniprot