Recombinant Human Transglutaminase 3/TGM3 Protein, CF

Catalog # Availability Size / Price Qty
4604-TG-050
R&D Systems Recombinant Proteins and Enzymes
1 Image
Product Details
FAQs
Supplemental Products
Reviews

Recombinant Human Transglutaminase 3/TGM3 Protein, CF Summary

Product Specifications

Purity
>95%, by SDS-PAGE under reducing conditions and visualized by silver stain.
Endotoxin Level
<1.0 EU per 1 μg of the protein by the LAL method.
Activity
Measured by its ability to cleave a synthetic peptide Benzyloxycarbonyl-Gln-Gly and NH2OH. The specific activity is >450 pmol/min/µg, as measured under the described conditions.
Source
Spodoptera frugiperda, Sf 21 (baculovirus)-derived human Transglutaminase 3/TGM3 protein
Ala2-Glu693 (Gly654Arg), with an N-terminal Met and 6-His tag
Accession #
N-terminal Sequence
Analysis
No result obtained
Predicted Molecular Mass
72 kDa
SDS-PAGE
72 kDa, reducing conditions

Product Datasheets

You must select a language.

x

4604-TG

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.

4604-TG

Formulation Supplied as a 0.2 μm filtered solution in Tris, NaCl and Glycerol.
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.

Assay Procedure

Materials
  • Assay Buffer: 50 mM Tris, 10 mM CaCl2, 0.15 M NaCl, 0.05% Brij, pH 7.5 (TCNB)
  • Assay Diluent: Deionized water
  • Recombinant Human Transglutaminase 3/TGM3 (rhTGM3) (Catalog # 4604-TG)
  • Bacterial Thermolysin (Thermolysin) (Catalog # 3097-ZN)
  • Substrate: Z-Gln-Gly (Sigma, Catalog # C6154), dissolve 500 mM in deionized water, then adjust to pH 9.0 with NaOH
  • MES, pH 6.0, 400 mM stock in deionized water
  • DTT (Sigma, Catalog # D-0632), 200 mM stock in deionized water
  • CaCl2, 200 mM stock in deionized water
  • 1 M Hydroxylamine Hydrochloride (Sigma, Catalog # 159417), dissolve in deionized water, then adjust to pH 6.0 with NaOH
  • Stop Solution: 0.37 M FeCl3 (Sigma, Catalog # 236489), 0.67 M HCl, 0.2 M Trichloroacetic Acid
  • 96-well Clear Plate (Costar, Catalog # 92592)
  • Plate Reader (Model: Spectramax Plus by Molecular Devices) or equivalent
  1. Combine the following volumes per reaction vial: 10 µL of 500 mM Substrate, 50 µL of 400 mM MES, pH 6.0, 5 µL of 200 mM DTT, 5 µL of 200 mM CaCl2, and 10 µL of 1.0 M Hydroxylamine Hydrochloride right before running assays. Note: Multiply the volume for each component by the number of reaction vials + 1 to make enough substrate mixture for the assays. For example, if there are 2 reaction vials (including blank), multiply all volumes by 3.
  2. Dilute rhTGM3 to 0.2 mg/mL with 2 ng/µL Thermolysin in Assay Buffer.
  3. Incubate 30 minutes at room temperature.
  4. Dilute rhTGM3 to 0.05 mg/mL in Assay Diluent.
  5. Mix 20 µL of the diluted rhTGM3 with 80 µL Substrate Mixture. For the Blank mix 20 µL of Assay Diluent with 80 µL Substrate Mixture.
  6. Incubate at 37 °C for 2 hours.
  7. After incubation, stop the reaction with 400 µL of the Stop Solution. Mix well.
  8. Centrifuge at top speed for 2 minutes in a microcentrifuge.
  9. Load 200 µL of the supernatant into a plate.
  10. Read at 525 nm (absorbance) in endpoint mode.
  11. Calculate specific activity:

     Specific Activity (pmol/min/µg) =

Adjusted Abs* (OD) x Conversion Factor** (pmol/OD)
Incubation time (min) x amount of enzyme (µg)

     *Adjusted for Substrate Blank

     **Derived using calibration standard L-glutamic acid g-monohydroxamate (Sigma, Catalog # G2253).

Per Well:
  • rhTGM3: 0.4 µg
  • Substrate: 10 mM
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.

=
÷

Background: Transglutaminase 3/TGM3

Transglutaminase 3 (TG3), also known as epidermal Transglutaminase (Tgase E), belongs to the family of Transglutaminase enzymes that catalyze the posttranslational modification of proteins via calcium dependent cross-linking reactions (1-3). TG3 is involved in the formation of the cornified envelope in skin keratinocytes (4). It functions to cross-link structural proteins during epidermal terminal differentiation. TG3 has been implicated as the dominant autoantigen in dermatitis herpetiformis (5). TG3 activation requires proteolysis of the 77 kDa zymogen into two fragments of approximately 50 and 27 kDa to form the active enzyme (1).

References
  1. Kim, I.G. et al. (1993) J. Biol. Chem. 268:12682.
  2. Griffin, M. et al. (2002) Biochem. J. 368:377.
  3. Lorand, L. and R.M. Graham (2003) Nat. Rev. Mol. Cell Biol. 4:140.
  4. Eckert, R.L. et al. (2005) J. Invest. Dermatol. 124:481.
  5. Sardy, M. et al. (2002) J. Exp. Med. 195:747.
Entrez Gene IDs
7053 (Human); 21818 (Mouse)
Alternate Names
EC 2.3.2.13; MGC126250; protein-glutamine gamma-glutamyltransferase E; TG(E); TG3; TGase E; TGase-3; TgaseE; TGEMGC126249; TGM3; transglutaminase 3 (E polypeptide, protein-glutamine-gamma-glutamyltransferase); Transglutaminase 3; Transglutaminase E; transglutaminase-3

FAQs

No product specific FAQs exist for this product, however you may

View all Proteins and Enzyme FAQs

Reviews for Recombinant Human Transglutaminase 3/TGM3 Protein, CF

There are currently no reviews for this product. Be the first to review Recombinant Human Transglutaminase 3/TGM3 Protein, CF and earn rewards!

Have you used Recombinant Human Transglutaminase 3/TGM3 Protein, CF?

Submit a review and receive an Amazon gift card.

$25/€18/£15/$25CAN/¥75 Yuan/¥1250 Yen for a review with an image

$10/€7/£6/$10 CAD/¥70 Yuan/¥1110 Yen for a review without an image

Submit a Review