GST-PAK-CRIB Rac/Cdc42 Isolation Kit
Recombinant protein purified from E. coli, consisting of GST fused to the Cdc42 (p21) binding domain (CRIB) of the human p21 activated kinase 1 protein (PAK).
Highlights:
- Specifically binds to GTP-bound (but not GDP-bound) Cdc42 small GTPase for precipitation of active, GTP-bound Cdc42 from cell lysates
- Contains the highly conserved CRIB motif plus sequences required for the high affinity interaction with GTP-Cdc42
- Provided as GST-PAK-CRIB non-covalently bound to glutathione agarose
This non-radioactive pulldown assay uses the p21 binding domain (PBD) of the Rac/Cdc42, p21-activated protein kinase (PAK1). This domain is expressed as an N-terminal GST-fusion protein coupled to glutathione beads and can be used to isolate the active, GTP-bound form of Rac/Cdc42 from whole cell lysates. After precipitation, Rac/Cdc42 can be detected by immunoblotting with the appropriate antibodies.
From the laboratory of Jorrit M. Enserink, PhD, University of Oslo.
Recombinant protein purified from E. coli, consisting of GST fused to the Cdc42 (p21) binding domain (CRIB) of the human p21 activated kinase 1 protein (PAK).
Highlights:
- Specifically binds to GTP-bound (but not GDP-bound) Cdc42 small GTPase for precipitation of active, GTP-bound Cdc42 from cell lysates
- Contains the highly conserved CRIB motif plus sequences required for the high affinity interaction with GTP-Cdc42
- Provided as GST-PAK-CRIB non-covalently bound to glutathione agarose
This non-radioactive pulldown assay uses the p21 binding domain (PBD) of the Rac/Cdc42, p21-activated protein kinase (PAK1). This domain is expressed as an N-terminal GST-fusion protein coupled to glutathione beads and can be used to isolate the active, GTP-bound form of Rac/Cdc42 from whole cell lysates. After precipitation, Rac/Cdc42 can be detected by immunoblotting with the appropriate antibodies.
From the laboratory of Jorrit M. Enserink, PhD, University of Oslo.
| Product Type: | Protein |
| Name: | GST-PAK-CRIB |
| Accession ID: | Q86VW2, Q15052 |
| Source: | E. coli |
| Molecular Weight: | 34 kDa GST fusion protein |
| Amino Acid Sequence: | RSILPGDKTNKKKEKERPEISLPSDFEHTIHVGFDAVTGEFTGMPEQWARLLQTSNITKSEQKKNPQAVLDVLEFYNSKKTSNSQKYMSFTDKSA |
| Fusion Tag(s): | GST |
| Purity: | >90% |
| Buffer: | 30% glycerol, PBS pH7.4, 2 mM EDTA, 10 mM DTT, protease inhibitors |
| Concentration: | Amount of product necessary to pull down active GTPase in the linear range is between 2-2.5uL (recommended amount per assayed sample - see "DOCUMENTATION" section) |
| Suggested Amount per Experiment: | 2-2.5uL per assays sample |
| Storage: | -80C |
| Shipped: | Dry ice |
Protocol Information
Recommended amount of beads per assayed sample: 2-2.5 µl (higher amounts are unnecessary and may even negatively affect experimental outcome).
Protein not quantified as this would not provide information on its actual activity.
- Benard V, Bohl BP, Bokoch GM. Characterization of rac and cdc42 activation in chemoattractant-stimulated human neutrophils using a novel assay for active GTPases. J Biol Chem. 1999 May 7;274(19):13198-204.
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