ANTIBODIES
Contributor Information
- Name Brian Burke
- Institute A*STAR Accelerate Technologies Pte Ltd
Tool Details
- Tool name: AntiBirA [5B11c3-3]
- Alternate names: BirA antibody, bioR antibody, dhbB antibody, JW3941 antibody, BirA miniTurbo antibody, BirA/miniTurbo
- Clone: 5B11c3-3
- Tool type: Antibodies
- Tool sub-type: Primary Antibody
- Class: Monoclonal
- Conjugate: Unconjugated
- Reactivity: E.coli
- Host: Mouse
- Molecular weight of the target: 35 kDa
- Application: IF ; WB
- Strain: C57BL/6
- Description: Monoclonal antibody which specifically detects BirA in E.coli samples. Background and Research Application E.coli BirA is a well-characterized multifunctional protein that either catalyses the transfer of biotin to biotin carboxyl carrier protein (BCCP), an acetyl-CoA carboxylase subunit or, alternatively, binds to the biotin operator resulting in transcriptional repression of the biotin operon. This protein also activates biotin to form biotinyl-5'-adenylate and transfers the biotin moiety to biotin-accepting proteins. A mutant BirA (R118G) shows loss of DNA binding ability and promiscuity of substrate biotinylation. These properties of BirA R118G were used to develop an in vivo proximity labelling technique that identifies potential interacting proteins (BioID). In the BioID method, a protein of interest is tagged with BirA R118G and expressed in live cells. Addition of the biotin to the culture medium results in biotinylation of proteins in vicinity of the bait. The biotinylated proteins can then be affinity purified with biotin binding proteins such as streptavidin or avidin even under harsh denaturing conditions and subsequently identified by mass spectrometry or immunoblot analysis. One of the drawbacks of the BirA/BioID is its slow biotinylation kinetics making it unsuitable for the study of dynamic processes. To overcome the hurdle directed evolution of BirA R118G was used to generate two BirA variants with superior enzymatic activity: (i) 35 kD TurboID, with 15 mutations relative to wild-type BirA; and 28 kD miniTurbo, with N-terminal domain deleted and 13 mutations relative to the original BirA. This antibody specifically detects BirA in E. coli samples.
- Immunogen: Glutathione-S-transferase (GST) fused to E.coli BirA R118G
- Immunogen UniProt ID: P06709
- Isotype: IgG1 kappa
- Research area: Epitope Tags
- Myeloma used: Sp2/0-Ag14
- For Research Use Only
Target Details
- Target: BirA R118G
- Target molecular weight: 35 kDa
- Target background: Monoclonal antibody which specifically detects BirA in E.coli samples. Background and Research ApplicationE.coli BirA is a well-characterized multiFn protein that either catalyses the transfer of biotin to biotin carboxyl carrier protein (BCCP), an acetyl-CoA carboxylase subunit or, alternatively, binds to the biotin operator resulting in transcriptional repression of the biotin operon. This protein also activates biotin to form biotinyl-5'-adenylate and transfers the biotin moiety to biotin-accepting proteins. A mutant BirA (R118G) shows loss of DNA binding ability and promiscuity of substrate biotinylation. These properties of BirA R118G were used to develop an in vivo proximity labelling technique that identifies potential interacting proteins (BioID). In the BioID method, a protein of interest is tagged with BirA R118G and expressed in live cells. Addition of the biotin to the culture medium results in biotinylation of proteins in vicinity of the bait. The biotinylated proteins can then be affinity purified with biotin binding proteins such as streptavidin or avidin even under harsh denaturing conditions and subsequently identified by mass spectrometry or immunoblot analysis.One of the drawbacks of the BirA/BioID is its slow biotinylation kinetics making it unsuitable for the study of dynamic processes. To overcome the hurdle directed evolution of BirA R118G was used to generate two BirA variants with superior enzymatic activity: (i) 35 kD TurboID, with 15 mutations relative to wild-type BirA; and 28 kD miniTurbo, with N-terminal domain deleted and 13 mutations relative to the original BirA. This antibody specifically detects BirA in E. coli samples.
Application Details
- Application: IF ; WB
Handling
- Format: Liquid
- Concentration: 1 mg/ml
- Storage buffer: PBS with 0.02% azide
- Storage conditions: Store at -20°C frozen. Avoid repeated freeze / thaw cycles
- Shipping conditions: Shipping at 4°C
Documentation
- Available on request
References
- • Creed et al. 2020. Development. 147(1):. PMID: 31806659.
- • Branon et al. 2018. Nat Biotechnol. 36(9):880-887. PMID: 30125270.
- • Roux et al. 2012. J Cell Biol. 196(6):801-10. PMID: 22412018.
- • Choi-Rhee et al. 2004. Protein Sci. 13(11):3043-50. PMID: 15459338.
- • Streaker et al. 2003. J Mol Biol. 325(5):937-48. PMID: 12527300.
- • Kwon et al. 2000. J Mol Biol. 304(5):821-33. PMID: 11124029.
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