Human Transglutaminase 3/TGM3 Baculovirus-Insect cells Overexpression Lysate: Product Information
This Human Transglutaminase 3/TGM3 overexpression lysate was created in Baculovirus-Insect cells and intented for use as a Western blot (WB) positive control. Purification of Transglutaminase 3/TGM3 protein (Cat: 11923-H07B) from the overexpression lysate was verified.
A DNA sequence encoding the human TGM3 (Q08188) (Ala 2-Glu 693) was expressed, with a polyhistidine tag at the N-terminus.
The recombinant human TGM3 consists of 710 amino acids and predicts a molecular mass of 78.8 kDa. It migrates as an approximately 70 kDa band in SDS-PAGE under reducing conditions.
Human Transglutaminase 3/TGM3 Baculovirus-Insect cells Overexpression Lysate: Usage Guide
Cell lysate was prepared by homogenization of the over-expressed cells in ice-cold modified RIPA Lysis Buffer with cocktail of protease inhibitors (Sigma). Cell debris was removed by centrifugation. Protein concentration was determined by Bradford assay (Bio-Rad protein assay, Microplate Standard assay). The cell lysate was boiled for 5 min in 1 x SDS loading buffer (50 mM Tris-HCl pH 6.8, 12.5% glycerol, 1% sodium dodecylsulfate, 0.01% bromophenol blue) containing 5% b-mercaptoethanol, and lyophilized.
1. Centrifuge the tube for a few seconds and ensure the pellet at the bottom of the tube.
2. Re-dissolve the pellet using 200μL pure water and boil for 2-5 min.
1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).
Stability & Storage
Store at 4℃ for up to twelve months from date of receipt. After re-dissolution, aliquot and store at -80℃ for up to twelve months. Avoid repeated freeze-thaw cycles.
Western Blot (WB) Optimal dilutions/concentrations should be determined by the end user.
Human Transglutaminase 3/TGM3 Baculovirus-Insect cells Overexpression Lysate: Alternative Names
Human TGE Overexpression Lysate; Human UHS2 Overexpression Lysate
Transglutaminase 3/TGM3 Background Information
Transglutaminases (TGase) are a family of calcium-dependent acyl-transfer enzymes ubiquitously expressed in mammalian cells and responsible for catalyzing covalent cross-links between proteins or peptides. Transglutaminase 3 (TGM3) is a member of a family of Ca2+-dependent enzymes that catalyze covalent cross-linking reactions between proteins or peptides. TGM3 isoform is widely expressed and is important for epithelial barrier formation. It is a zymogen, requiring proteolysis for activity. Calcium-activated TGM3 can bind, hydrolyze, and is inhibited by GTP, despite lacking structural homology with other GTP binding proteins. TGM3 displays a diffuse cytoplasmic distribution in vitro consistent with its proposed role in the early phase of cornified cell envelope assembly in the cytoplasm. TGM3-driven specific isopeptide bonds between intermediate filaments and KAPs participate to the progressive scaffolding of the hair shaft. Additionally, TGM3 may be a novel prognostic biomarker for esophageal squamous cell carcinoma (ESCC).
Ahvazi B, et al. (2002) Three-dimensional structure of the human transglutaminase 3 enzyme: binding of calcium ions changes structure for activation. EMBO J. 21(9): 2055-67.
Hitomi K, et al. (2003) Analysis of epidermal-type transglutaminase (transglutaminase 3) in human stratified epithelia and cultured keratinocytes using monoclonal antibodies. J Dermatol Sci. 32(2): 95-103.
Ahvazi B, et al. (2004) The emerging structural understanding of transglutaminase 3. J Struct Biol. 147(2): 200-7.
Ahvazi B, et al. (2004) Structural basis for the coordinated regulation of transglutaminase 3 by guanine nucleotides and calcium/magnesium. J Biol Chem. 279(8): 7180-92.
Uemura N, et al. (2009) Transglutaminase 3 as a prognostic biomarker in esophageal cancer revealed by proteomics. Int J Cancer. 124(9): 2106-15.
Thibaut S, et al. (2009) Transglutaminase-3 enzyme: a putative actor in human hair shaft scaffolding? J Invest Dermatol. 129(2): 449-59.
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