Recombinant Human Transglutaminase 4/TGM4 Protein, CF

Catalog #: 5760-TG Datasheet / COA / SDS

Catalog #	Availability	Size / Price	Qty
5760-TG-050

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Recombinant Human Transglutaminase 4/TGM4 Protein, CF Summary

Product Specifications

Purity

>95%, by SDS-PAGE under reducing conditions and visualized by Colloidal Coomassie® Blue stain at 5 μg per lane

Endotoxin Level

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

Activity

Measured by its ability to form CBZ-Gln-Gly-Hydroxamate from CBZ-Gln-Gly and Hydroxylamine. The specific activity is >50 pmol/min/μg, as measured under the described conditions.

Source

Spodoptera frugiperda, Sf 21 (baculovirus)-derived human Transglutaminase 4/TGM4 protein
Met2-Lys684, with an N-terminal Met and 6-His tag

Accession #

AAC50516

N-terminal Sequence
Analysis

No results obtained, Met predicted. Protein identity confirmed by MS analysis of tryptic fragments.

Predicted Molecular Mass

78 kDa

SDS-PAGE

65-70 kDa, reducing conditions

Product Datasheets

5760-TG

Product Datasheet

COA

SDS

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.

5760-TG

Formulation	Supplied as a 0.2 μm filtered solution in Tris, NaCl, DTT and Glycerol.
Shipping	The product is shipped with dry ice or equivalent. 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.

Assay Procedure

Materials

Assay Buffer: deionized water
Recombinant Human Transglutaminase 4/TGM4 (rhTGM4) (Catalog # 5760-TG)
Substrate: Z-Gln-Gly (Sigma, Catalog # C6154), 500 mM, pH 9.0 in deionized water
Hydroxylamine Hydrochloride (Sigma, Catalog # 159417), 1 M, pH 6.0 in deionized water
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

Prepare Substrate Mixture containing 62.5 mM substrate, 250 mM sodium acetate pH 5.0, 12.5 mM DTT, 200 mM CaCl₂, 125 mM hydroxylamine hydrochloride, and 627 mM NaCl in deionized water.
Dilute rhTGM4 to 0.2 mg/mL in Assay Buffer.
Mix 30 µL dilute rhTGM4 with 120 µL Substrate Mixture. Include an enzyme blank containing 30 µL Assay Buffer with 120 µL Substrate Mixture.
Incubate at 37 °C for 2 hours.
Stop each reaction with 600 µL of Stop Solution. Mix well.
Centrifuge at top speed for 2 minutes in a microcentrifuge.
Load 200 µL of the supernatant into a 96 well clear plate.
Read plate at 525 nm (absorbance) in endpoint mode.
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 gamma -monohydroxamate (Sigma, Catalog # G2253).

Per Well:

rhTGM4: 1.6 µg
Substrate: 10 mM
Hydroxylamine: 20 mM

Reconstitution Calculator

Background: Transglutaminase 4/TGM4

Transglutaminase 4, also known as TGM4, TG4, hTGP, or the prostate-specific transglutaminase, is a member of the transglutaminase family that is exclusively expressed and secreted from the prostate (1, 2). Transglutaminases catalyze the calcium-dependent formation of isopeptide bonds between lysine and glutamine residues (3). In rodents, TGM4 is responsible for the formation of the copulatory plug and sperm antigenicity, but the function in humans is largely unknown (4). The expression of TGM4 is primarily regulated by retinoic acid and secondarily modulated by androgens (5).

References

Dubbink, H.J. et al. (1996) Biochem. J. 315:901.
Dubbink, H.J. et al. (1999) Lab. Invest. 79:141.
Lorand, L. and Graham, R.M. (2003) Nature Rev. Mol. Cell. Biol. 4:140.
Williams-Ashman, H.G. et al. (1972) Proc. Natl. Acad. Sci. USA 69:2322.
Rivera-Gonzalez, G.C. et al. (2012) Nucleic Acids Res. 40:4825.

Entrez Gene IDs

7047 (Human); 331046 (Mouse); 64679 (Rat)

Alternate Names

EC 2.3.2.13; Fibrinoligase; hTGP; Prostate transglutaminase; Prostate-specific transglutaminase; protein-glutamine gamma-glutamyltransferase 4; TG(P); TGase P; TGase-4; TGM4; TGP; TGPFLJ26776; transglutaminase 4 (prostate); Transglutaminase 4; Transglutaminase P; transglutaminase-4