Cdc42-His Protein: constituitively active

Question 1: Does the constitutively-active mutant Cdc42 protein come pre-loaded with GTP?

Answer 1: The purified small G-proteins that Cytoskeleton provides, whether they are wild-type, constitutively-active or dominant negative proteins, do not come pre-loaded with GTP. Typically, 50% of the protein is bound to GTP and 50% is bound to GDP upon resuspension. Thus, proteins will have to be pre-loaded with either GTP (Cat. #BST06) or GTPgS (Cat. #BS01) for experiments.

Question 2: In what buffer is the mutant Cdc42 protein shipped?

Answer 2: The His-Cdc42 L61 mutant protein ((Cat. #C6101) is supplied as a lyophilized white powder. The protein should be reconstituted to 1 mg/ml by the addition of 10 &mu, l of distilled water. The protein will be in the following buffer: 10 mM Tris pH 7.5, 10 mM NaCl, 0.1 mM MgCl₂, 0.5% sucrose and 0.1% dextran. In order to maintain high biological activity of the protein, it is strongly recommended that the protein solution be supplemented with DTT to 1 mM final concentration, aliquoted into "experiment-sized" amounts, snap frozen in liquid nitrogen and stored at -70°C. The protein is stable for 6 months under these conditions. The protein must not be exposed to repeated freeze-thaw cycles.