Product Uses Include

      Stimulating Arp2/3 complex activity

      Isolating Arp2/3 complex from cell or tissue extract

Material
     The VCA (Verprolin, cofilin, acidic) domain of human WASP protein has been expressed in a bacterial expression system as a GST-tagged fusion protein. The protein has been purified by glutathione affinity chromatography and is supplied as a lyophilized powder. When reconstituted with distilled water, the complex is in the following buffer: 20 mM Tris pH 7.5, 25 mM KCl, 1 mM MgCl2, 0.5 mM EDTA, 0.2% dextran, 2% sucrose. The molecular weight of the GST-VCA domain protein is approximately  43 kDa.

Purity
     Purity is determined by scanning densitometry of proteins on SDS-PAGE gels. Samples are >95% pure (Figure 1).

Figure 1: Purity determination of GST-WASP VCA domain. 20 μg of VCG03 was run on an SDS-PAGE gel and visualized by coomassie blue staining. Protein quantitation was performed using the Precision Red Protein Assay Reagent (Cat.# ADV02).

Biological Activity
      VCG03 was tested in an actin polymerization assay (Cat. # BK003). In conjunction with Arp2/3 (Cat. # RP01) it was shown to stimulate actin polymerization 20-fold compared to the control without VCG03. This indicates VCA domain specificity for stimulating the actin nucleating activity of Arp2/3.

Figure 2: Actin polymerization stimulated by Arp2/3 complex and the VCA domain of WASP. Actin polymerization was measured using kit BK003. The addition of Arp2/3 complex or the VCA domain alone to actin has minimal effects on actin polymerization, while the combination of Arp2/3 and the VCA domain strongly stimulates the rate of actin polymerization.