Recombinant Human C1GalT1/C1GalT1C1 Protein, CF

R&D Systems | Catalog # 8659-GT

R&D Systems
100% Guarantee

Key Product Details

  • R&D Systems CHO-derived Recombinant Human C1GalT1/C1GalT1C1 Protein (8659-GT)
  • 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

Q9NS00 (C1GalT1) & Q96EU7 (C1GalT1C1)

Applications

Enzyme Activity
View all C1GalT1 Proteins and Enzymes »

Product Specifications

Source

Chinese Hamster Ovary cell line, CHO-derived human C1GalT1 protein
Asp45-Pro363, with N-terminal 6-His tag (C1GalT1); Ile28-Asp318, with an N-terminal HA tag (C1GalT1C1)

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

His (C1GalT1) & Tyr (C1GalT1C1)

Predicted Molecular Mass

38 kDa (C1GalT1) & 35 kDa (C1GalT1C1)

SDS-PAGE

25-42 kDa, reducing conditions

Activity

Measured by its ability to transfer galactose from UDP-galactose to 4-nitrophenyl-alpha -D-galactosaminide.
The specific activity is >2,750 pmol/min/μg, as measured under the described conditions.

Formulation, Preparation, and Storage

8659-GT
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: C1GalT1

O-glycosylation is a ubiquitous post-translational modification of secreted and membrane bound proteins (1). The synthesis of mucin-type O-glycans is initiated by the addition of GalNAc to threonine or serine residues on proteins by polypeptide N-acetylgalactosaminyltransferases (GALNTs) (2). The GalNAc alpha 1-O-Ser/Thr structure is then extended by other glycosyltransferases to form eight types of core O-glycans (3). Core 1 beta -3-galactosyltransferase (C1GalT1), in particular, synthesizes Gal-beta 1-3GalNAc alpha 1-O-Ser/Thr, a precursor for all core 1 and core 2 based mucin-type O-glycans (4). These glycans play central roles in many processes, such as angiogenesis, thrombopoiesis, and kidney homeostasis (5). C1GalT1 forms a stable, non-covalent complex with Cosmc chaperone, C1GalT1C1, which is required for the full activity of C1GalT1 (6). Defective C1GalT1 causes a rare autoimmune disease called Tn syndrome (4) as well as susceptibility to IgA nephropathy (7). The recombinant C1GalT1 is co-purified with C1GalT1C1. The enzymatic activity of recombinant human C1GalT1 was determined using a phosphatase-coupled assay (8).

References

  1. Gerken, T.A. et al. (2011) J. Biol. Chem. 286:14493.
  2. Bergstrom, K.S.B. and Xia, L. (2013) Glycobiology 23:1026.
  3. Brockhausen, I. et al. (1999) Biochim. Biophys. Acta. 1473:67.
  4. Ju, T. and Cummings, R.D. (2005) Nature 437:1252.
  5. Fukuda, M. et al. (2002) Biochim. Biophys. Acta. 1573:394.
  6. Aryal, R.P. et al. (2012) J. Biol. Chem. 287:15317.
  7. Pirulli, D. et al. (2009) J. Nephrol. 22:152.
  8. Wu, Z.L. et al. (2011) Glycobiology 21:727.

Long Name

Core 1 Synthase, Glycoprotein-N-Acetylgalactosamine 3-Beta-Galactosyltransferase, 1

Alternate Names

B3Gal-T8, Core 1 Beta3-Gal-T1, Core 1 O-Glycan T-Synthase

Entrez Gene IDs

56913 (Human); 94192 (Mouse); 65044 (Rat)

Gene Symbol

C1GALT1

UniProt

Q9NS00 (C1GalT1) & Q96EU7 (C1GalT1C1)

Additional C1GalT1 Products

Product Documents for Recombinant Human C1GalT1/C1GalT1C1 Protein, CF

Certificate of Analysis

To download a Certificate of Analysis, please enter a lot or batch number in the search box below.

Note: Certificate of Analysis not available for kit components.

Download SDS

Product Specific Notices for Recombinant Human C1GalT1/C1GalT1C1 Protein, CF

For research use only

Related Research Areas

Glycobiology-related Enzymes

Citations for Recombinant Human C1GalT1/C1GalT1C1 Protein, CF

Customer Reviews for Recombinant Human C1GalT1/C1GalT1C1 Protein, CF

There are currently no reviews for this product. Be the first to review Recombinant Human C1GalT1/C1GalT1C1 Protein, CF and earn rewards!

Have you used Recombinant Human C1GalT1/C1GalT1C1 Protein, CF?

Submit a review and receive an Amazon gift card!

$25/€18/£15/$25CAN/¥2500 Yen for a review with an image

$10/€7/£6/$10CAN/¥1110 Yen for a review without an image

Submit a review
Amazon Gift Card

Protocols

View specific protocols for Recombinant Human C1GalT1/C1GalT1C1 Protein, CF (8659-GT):

Materials
  • Glycosyltransferase Activity Kit (Catalog # EA001)
  • Assay Buffer: 25 mM Tris, 5 mM CaCl2, 10 mM MnCl2, pH 7.5
  • Recombinant Human C1GalT1 (rhC1GalT1) (Catalog # 8659-GT)
  • Uridine 5'-diphosphogalactose (UDP-Gal) (Sigma, Catalog # U4500), 10 mM stock in deionized water
  • 4-Nitrophenyl N-acetyl-alpha -D-galactosaminide (4-NP-GalNAc) (Sigma, Catalog # N4264), 15 mM stock in DMSO
  • 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 Glycosyltransferase 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. Prepare reaction mixture containing 1.2 mM UDP-Gal, 0.6 mM 4-NP-GalNAc, and 4 µg/mL Coupling Phosphatase 1 in Assay Buffer.
  4. Dilute rhC1GALT1 to 0.6 µg/mL 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 25 µL of 0.6 µg/mL rhC1GALT1 into empty wells of the same plate as the curve. Include a Control containing 25 μL of Assay Buffer.
  7. Add 25 µL of the reaction mixture to all wells, excluding the standard curve.
  8. Seal plate and incubate at 37 °C for 20 minutes.
  9. Add 30 µL of the Malachite Green Reagent A to all wells. Mix briefly.
  10. Add 100 µL of deionized water to all wells. Mix briefly.
  11. Add 30 µL of the Malachite Green Reagent B to all wells. Mix and incubate sealed plate for 20 minutes at room temperature.
  12. Read plate at 620 nm (absorbance) in endpoint mode.
  13. 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. Per Well:

  • rhC1GALT1: 0.015 µg
  • Coupling Phosphatase 1: 0.1 µg
  • UDP-Gal: 0.6 mM
  • 4-NP-GalNAc: 0.3 mM

FAQs

No product specific FAQs exist for this product.

View all FAQs for Proteins and Enzymes