Active Rac-GEF Assay Kit

Background: Small GTP-binding proteins (or GTPases) are a family of proteins that serve as molecular regulators in signal transduction pathways. Rac, a 21 kDa protein, belongs to the family of Rho GTPases regulating a variety of biological response pathways that include cell motility, cell division, gene transcription, and cell transformation. Like other small GTPases, Rac influences molecular events by cycling between an inactive GDP-bound form and an active GTP-bound form. Cycling between the GDP-bound and GTP-bound state is regulated primarily by two distinct families of proteins: guanine nucleotide exchange factors (GEFs) activate Rho proteins by catalyzing the exchange of GDP for GTP, the GTPase activating proteins or GAPs negatively regulate GTPase function by stimulating GTP hydrolysis (Figure 1). Similar to Ras mutants, constitutively active or dominant negative Rho GTPase mutants have been used to bind to Rho-GAP and effectors or to Rho-GEFs, respectively. A nucleotide-free GTPase has also been shown to form a high affinity binary complex with Rho-GEFs. Active Rac-GEF Assay Kit utilizes nucleotide-free Rac1 G15A Agarose beads to selectively isolate and pull-down the active form of Rac-GEFs from purified samples or endogenous lysates. Subsequently, the precipitated active Rac-GEF is detected by western blot analysis using an anti-Rac-GEF antibody (see Assay Principle below). Active Rac-GEF Assay Kit provides a simple and fast tool to monitor the activation of Rac-GEF. The kit includes easily identifiable Rac1 G15A Agarose beads (Figure 2), pink in color and anti-Tiam1 antibody. Each kit provides sufficient quantities to perform 20 assays.

1. Arthur, W.T., Ellerbroek, S.M., Der, C.J., and Burridge K. (2002) J. Biol. Chem. 277, 42964-42972
2. Garcia-Mata R., Wennerberg, K., Arthur, W.T., Noren, N.K., Ellenbroek, S.M., and Burridge K. (2006) Methods Enzymol. 406, 425-437.
3. Habets, G.G., Scholtes, E.H., Zuydgeest, D., van der Kammen, R.A., Stam, J.C., Berns, A. and Collard, J.G. (1994) Cell. 77, 537-549.
1. Oubaha, M. et al. (2012). Formation of a PKC/beta-Catenin Complex in Endothelial Cells Promotes Angiopoietin-1-Induced Collective Directional Migration and Angiogenic Sprouting. Blood. 120: 3371-3381.

Small volumes of Active Rac-GEF assay kit vial(s) may occasionally become entrapped in the seal of the product vial during shipment and storage. If necessary, briefly centrifuge the vial on a tabletop centrifuge to dislodge any liquid in the container`s cap. Certain products may require to ship with dry ice and additional dry ice fee may apply.

MBS169137 is an assay kit targeting Active Rac-GEF. Also known as Active Rac-GEF Assay Kit (Tiam1).