Racgap1

Manufactured by Santa Cruz Biotechnology

RacGAP1 is a GTPase-activating protein (GAP) that regulates the activity of the Rac1 GTPase. RacGAP1 functions to increase the intrinsic GTPase activity of Rac1, thereby inactivating the Rac1 protein. This modulation of Rac1 signaling is important for various cellular processes.

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Lab products found in correlation

2 protocols using racgap1

Western Blot Analysis of Signaling Proteins

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Whole cell extracts of cultured cells were prepared in RIPA lysis buffer supplemented with phosphatase and protease inhibitors, and separated on SDS-PAGE gel. The following antibodies and dilution factors were used: phospho-eIF4E (1:1000, Cell Signaling), total eIF4E (1:1000, Santa Cruz Biotechnology), phospho-MNK1 (1:1000, Cell Signaling), total MNK1 (1:1000, Cell Signaling), phospho-ERK1/2 (1:1000, Cell Signaling), total ERK1/2 (1:2000, Santa Cruz), BRD4 (1:1000, Abcam), Rac1 (1:2000, EMD Millipore), RacGAP1 (1:2000, Santa Cruz), and HSP90 (1:3000, Santa Cruz). Blocking agent was 5% bovine serum albumin (BSA). Secondary anti-mouse IgG (A4416) and anti-rabbit IgG (A6667) antibodies were purchased from Sigma and used at a 1:3000 dilution factor. When necessary, membrane was stripped using Restore Western Blot Stripping Buffer (Thermo Fisher Scientific).

Pham T.N., Kumar K., DeCant B.T., Shang M., Munshi S.Z., Matsangou M., Ebine K, & Munshi H.G. (2018). Induction of MNK kinases-dependent eIF4E phosphorylation by inhibitors targeting BET proteins limits efficacy of BET inhibitors. Molecular cancer therapeutics, 18(2), 235-244.

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Protein Quantification and Immunoblotting

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Protein samples were quantified by 1D SDS-PAGE/SYPRO Ruby protein staining-based densitometry^{18 (link)}. Western blotting was performed on protein samples (10–20 µg) as previously described^{18 (link)}. Rabbit antibodies raised against GAPDH (Cell Signalling), β-tubulin (Cell Signalling), KRAS^G12V mutant specific (Cell Signalling), RAB7 (Abcam) and GFP (Abcam) were used. Mouse antibodies against MKLP1 (Santa Cruz), RACGAP1 (Santa Cruz), ALIX (BD Biosciences), TSG101 (BD Biosciences), CD63 (Santa Cruz), CD81 (Santa Cruz), RAB2A (Thermo Fisher), FLOT1 (BD Biosciences), α-actinin (Abcam), CD9 (Santa Cruz) and HSP90 (BD Biosciences) were used. Secondary antibodies used were IRDye 800 goat anti-mouse IgG or IRDye 700 goat anti-rabbit IgG (1:15000, LI-COR Biosciences).

Rai A., Greening D.W., Xu R., Chen M., Suwakulsiri W, & Simpson R.J. (2021). Secreted midbody remnants are a class of extracellular vesicles molecularly distinct from exosomes and microparticles. Communications Biology, 4, 400.

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