K-Ras4B Mutant Protein: G13D (human recombinant with 6xHis-tag)

On Arrival: 4°C
Before reconstitution, briefly centrifuge to collect the product at the bottom of the tube. The protein should be reconstituted to 5 mg/ml with the addition of 20 ?l of Milli-Q water (100 ?g size).When reconstituted, the protein will be in the following buffer: 50 mM Tris pH 7.5, 50 mM NaCl, 0.5 mM MgCl₂ 5% (w/v) sucrose, and 1% (w/v) 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 -70C. The protein is stable for six months if stored at - 70C. The protein should not be exposed to repeated freezethaw cycles. The lyophilized protein is stable at 4C desiccated (< 10% humidity) for one year.

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The G13D (glycine to aspartic acid at amino acid position 13) mutant human K-Ras4B protein has been produced in a bacterial expression system. The recombinant protein contains six histidine residues at its amino terminus (His-tag). The molecular weight of 6xHis tagged G13D K-Ras4B is approximately 25 kDa and it is supplied as a white lyophilized powder.

1. Recombinant G13D K-Ras4B protein (Cat.# CS-RS06)
2. Recombinant SOS1-ExD protein (Cat.# CS-SOS1)
3. 2X Exchange Buffer (40 mM Tris pH 7.5, 100 mM NaCl, 20mM MgCl₂ 0.1 mg/ml BSA, 1.5 ?M mant-GTP)
4. Dilution Buffer (20 mM Tris pH 7.5, 50 mM NaCl, 10 mM MgCl₂ 0.1 mg/ml BSA)

1. Fluorescence spectrophotometer (?ex=360 nm, ?em=440 nm)
2. Corning 96-well half area plates (Cat. # 3686) or other plate with low protein binding surface.

Figure 2. GTP-exchange assay with KRas4B G13D and SOS1. KRas4B G13D (Cat. # CS-RS06) was used in a nucleotide exchange reaction Bodipy-GTP from ThermoFisher was used as a reporter at 1 ?M and KRas was at 2 ?M. Ex/Em was 490/513nm. Well A2 = Buffer control, B2 = Nucleotide free KRas, and C2 = Nucleotide free KRas with 100 ?M GTP competitor.