H-Ras Protein, wild type 6xHis Tag

Product Uses Include

• Ras biochemistry
• Ras GTPase assays
• Ras nucleotide exchange assays
• Ras binding studies
  
  

Material
The human H-Ras protein has been produced in a bacterial expression system. The protein is supplied as a lyophilized powder. When it is reconstituted in distilled water to 1 mg/ml, the protein is in the following buffer: 2 mM Tris pH 7.6, 0.5 mM MgCl₂ 0.5% sucrose, 0.1% dextran. Protein concentration is determined by the Precision Red Advanced Protein Assay Reagent, Cat. # ADV02.

The recombinant protein is approximately 28 kDa, consisting of the H-Ras protein plus a histidine tag in the amino-terminus.

For other forms of Ras as well as many other purified small G-proteins, see our main small G-protein product page.

Purity
Purity is determined by scanning densitometry of proteins on SDS-PAGE gels. His-H-Ras samples are > 80% pure.

Figure 1: His-H-Ras protein purity determination. A 10 ug sample of RS01 (His-H-Ras molecular weight approx. 28 kDa) was separated by electrophoresis in a 12% SDS-PAGE system. The protein was stained with Coomassie Blue.

Biological Activity
The biological activity of His-H-Ras is determined from its ability to catalyze the exchange of GDP for GTP. EDTA is used to sequester magensium ions from the His-H-Ras protein, thereby stimulating nucleotide exchange activity. The RhoGEF exchange assay Biochem Kit (Cat. # BK100) is used to monitor the nucleotide exchange in His-H-Ras.

Figure 2: His-Ras exchange assay using BK100. His-H-Ras protein (1 uM) was mixed with exchange buffer and aliquoted to four wells of a 96-well half area plate. After 5 cycles of reading in a fluorimeter, EDTA was added to 40 mM in the test samples and Milli-Q water to the control samples. Reactions were monitored for 30 min.

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