Product Uses Include
The Ras binding domain (RBD) of the c-Raf kinase protein (Raf) protein has been expressed as a GST-fusion protein in E. coli. This protein binds binds specifically to GTP-bound, and not GDP-bound, Ras proteins. The domain can therefore be used to specifically precipitate active, GTP-bound Ras as well as to specifically block the activity of Ras in vitro and in vivo.
The protein is supplied in a glutathione agarose bound format and is shipped lyophilized. The beads are colored for ease of use. This product is used in our Ras activation assay Biochem Kit™ (Cat. # BK008). The GST-tagged Raf-RBD protein can be released from the beads by incubation with 10 mM reduced glutathione.
Figure 1. The brightly colored glutatione agarose beads in RF02 are easy to use.
Protein purity is determined by scanning densitometry of Coomassie Blue stained protein on a 12% SDS polyacrylamide gel. GST-Raf-RBD protein is >80% pure (see Figure 2).
Figure 2: GST-Raf-RBD protein purity determination. A 20 µg sample of RF02 was separated by electrophoresis in a 12% SDS-PAGE system and stained with Coomassie Blue. The GST-Raf-RBD protein runs at approximately 35 kDa.
Raf-RBD protein specifically recognizes and binds the active, GTP-bound, forms of Ras proteins. It has a much lower affinity for the inactive, GDP-bound, form Ras. When coupled to a colored glutathione sepharose matrix, the Raf-RBD protein beads become a convienent tool for assaying the activity of the Ras proteins. The quality control biological assay for Raf-RBD protein beads consists of a Ras protein pulldown frombovine brain extracts loaded with either GTPγS (Cat. # BS01) or GDP.
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|Genest, Mallory et al.||Upregulated flotillins and sphingosine kinase 2 derail AXL vesicular traffic to promote epithelial-mesenchymal transition||Journal of Cell Science||2022||ISSN 0021--9533|
|Gao, Pin et al.||Nogo-B receptor increases the resistance to tamoxifen in estrogen receptor-positive breast cancer cells||Breast Cancer Research||2018||ISSN 1465-542X|
|Malilas, W. et al.||Cancer upregulated gene 2, a novel oncogene, confers resistance to oncolytic vesicular stomatitis virus through STAT1-OASL2 signaling||Cancer gene therapy||2013||ISSN 1476--5500|
|Takeishi, Kazuki et al.||Diacylglycerol kinase alpha enhances hepatocellular carcinoma progression by activation of Ras-Raf-MEK-ERK pathway||Journal of hepatology||2012||ISSN 1600--0641|
|Zhang, Jie et al.||Protein Kinase C (PKC) βII Induces Cell Invasion through a Ras/Mek-, PKCι/Rac 1-dependent Signaling Pathway *||Journal of Biological Chemistry||2004||ISSN 0021--9258|
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