Recombinant Human FAM20C Protein, CF

R&D Systems | Catalog # 9265-FM

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Key Product Details

  • R&D Systems CHO-derived Recombinant Human FAM20C Protein (9265-FM)
  • 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

CHO

Accession Number

Q8IXL6

Applications

Enzyme Activity
View all FAM20C Proteins and Enzymes »

Product Specifications

Source

Chinese Hamster Ovary cell line, CHO-derived human FAM20C protein
Val63-Arg584
with a C-terminal 6-His tag

Purity

>90%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.

Endotoxin Level

<1.0 EU per 1 μg of the protein by the LAL method.

N-terminal Sequence Analysis

Val63 (pro-mature) and Asp93 (mature)

Predicted Molecular Mass

60 kDa

SDS-PAGE

70-82 kDa, reducing conditions

Activity

Measured by its ability to hydrolyze the 5’-phosphate groups from the substrate adenosine-5’-triphosphate (ATP). The orthophosphate product is measured by a Universal Kinase Activity Kit (Catalog # EA004).
The specific activity is >20 pmol/min/μg, as measured under the described conditions.

Formulation, Preparation, and Storage

9265-FM
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.
  • 6 months from date of receipt, -20 to -70 °C as supplied.
  • 3 months, -20 to -70 °C under sterile conditions after opening.

Background: FAM20C

Fam20C is a protein kinase dedicated to the phosphorylation of extracellular proteins, including caseins, and is localizes within the Golgi apparatus and also is secreted (1, 2). It phosphorylates the caseins as well as several other secreted proteins such as AMELX, AMTN, ENAM, SPP1 and the small integrin-binding ligand N-linked glycoproteins SIBLINGs implicated in biomineralization (3). Consequently, mutations in Fam20C cause an osteosclerotic bone dysplasia in humans known as Raine syndrome (4) and hypophosphatemic rickets (5). The discovery of the function of Fam20C has solved a 130-year-old mystery of the identity of the kinases that phosphorylate caseins and has shed light on several human disorders of biomineralization (6). In addition to its role in biomineralization, Fam20C also plays a role in lipid homeostasis, wound healing and cell migration and adhesion (1).

References

  1. Tagliabracci, V. S. et al. (2013) cell 161:1619.
  2. Tagliabracci, V. S. et al. (2012) Science 336:6085.
  3. Cui, j. et al. (2015) Elife 4:e06120
  4. Fradin M, et al. (2011) Clin. Genet. 80:177.
  5. Wang X, et al. (2012) PLoS Genet. 8(5):e1002708.
  6. Tagliabracci VS, et al. (2013) Trends Biochem. Sci. 38:121.

Long Name

Family with Sequence Similarity 20, Member C

Alternate Names

DMP4, GEF-CK, RNS

Entrez Gene IDs

56975 (Human); 80752 (Mouse); 304334 (Rat)

Gene Symbol

FAM20C

UniProt

Q8IXL6

Additional FAM20C Products

Product Documents for Recombinant Human FAM20C Protein, CF

Certificate of Analysis

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Note: Certificate of Analysis not available for kit components.

Download SDS

Product Specific Notices for Recombinant Human FAM20C Protein, CF

For research use only

Related Research Areas

Intracellular Kinases

Citations for Recombinant Human FAM20C Protein, CF

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Protocols

View specific protocols for Recombinant Human FAM20C Protein, CF (9265-FM):

Materials
  • Universal Kinase Activity Kit (Catalog # EA004)
  • Assay Buffer: 25 mM HEPES, 150 mM NaCl, 10 mM MnCl2, 10 mM CaCl2, pH 7.0
  • Recombinant Human FAM20C (rhFAM20C) (Catalog # 9265-FM)
  • 96-well Clear Plate (Catalog # DY990)
  • Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
  1. 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 Assay Buffer for a 100 µM stock. This is the first point of the standard curve.
  2. Complete the standard curve by performing six one-half serial dilutions of the 100 µM Phosphate stock using Assay Buffer. The standard curve has a range of 0.078 to 5 nmol per well.
  3. Dilute ATP (supplied in kit) to 0.4 mM in Assay Buffer.
  4. Dilute rhFAM20C to 33.34 ng/µL in Assay Buffer.
  5. Load 50 µL of each dilution of the standard curve into a plate. Include a curve blank containing 50 μL of Assay Buffer.
  6. Load 15 µL of 33.34 ng/µL rhFAM20C into empty wells of the same plate as the curve. Include a Control containing 15 μL of Assay Buffer.
  7. Add 25 µL of 0.4 mM ATP into all wells, excluding the standard curve. 
  8. Seal plate and incubate at 37 °C for 1 hour.
  9. Dilute Coupling Phosphatase 4 (supplied in kit) to 10 ng/µL in Assay Buffer.
  10. Add 10 µL of 10 ng/µL Coupling Phosphatase 4 to all wells, excluding the standard curve.
  11. Seal and incubate plate at room temperature for 5 minutes.
  12. Add 30 µL of the Malachite Green Reagent A to all wells. Mix briefly.
  13. Add 100 µL of deionized water to all wells. Mix briefly.
  14. Add 30 µL of the Malachite Green Reagent B to all wells. Mix and incubate sealed plate for 20 minutes at room temperature.
  15. Read plate at 620 nm (absorbance) in endpoint mode.
  16. 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.
     ** Based upon a 65 minute incubation time.  (No coupling rate constant.) 

Per Reaction:

  • rhFAM20C: 0.5 µg
  • Coupling Phosphatase 4: 0.1 µg
  • ATP: 0.2 mM

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