H-Ras protein: His tagged: human wild type

H-Ras protein: His tagged: human wild type
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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 µg 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 µM) 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.

For product Datasheets and MSDSs please click on the PDF links below.   For additional information, click on the FAQs tab above or contact our Technical Support department at tservice@cytoskeleton.com

Z. Surviladze et al. 2010. Identification of a small GTPase inhibitor using a high-throughput flow cytometry bead-based multiplex assay. J. Biomol. Screen. 15, 10-20.

 

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