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Sulfo-SBED Biotin Label Transfer Reagent and Kit


A powerful tool for tagging and discovering protein interaction partners.

Pierce Sulfo-SBED Biotin Label Transfer Reagent

The Thermo Scientific Pierce Sulfo-SBED Biotin Label Transfer Reagent is a multifunctional reagent for labeling a purified protein and then covalently transferring the attached biotin tag onto specific interactors of that protein.

Sulfo-SBED is the abbreviation for Sulfo-N-hydroxysuccinimidyl-2-(6-[biotinamido]-2-(p-azido benzamido)-hexanoamido) ethyl-1,3'-dithioproprionate. It is a heterobifunctional chemical crosslinker capable of covalently attaching to primary amines at one end and to nearly any protein functional group at the other end. Unlike typical crosslinkers, Sulfo-SBED also includes a biotin group and a cleavable disulfide spacer arm. Together these features allow one to sequentially crosslink interacting proteins and transfer the biotin affinity tag from one protein (i.e., a purified "bait" protein) to another (possibly unknown "prey" protein). Label Transfer is a powerful in vitro method for protein interaction discovery. A growing number of publications feature the use of Sulfo-SBED Biotin Label Transfer Reagent to identify previously unknown protein interaction binding partners and to more fully characterize the specific protein binding domains of other protein interactions.

Product Details:


Sulfo-N-hydroxysuccinimidyl-2-(6-[biotinamido]-2-(p-azido benzamido)-hexanoamido) ethyl-1,3'-dithioproprionate; Sulfo-SBED Biotin
Structural and functional properties of Sulfo-SBED for biotin label transfer:
  • Amine-reactive NHS-ester group – for labeling a purified "bait" protein at the N-terminus and side chain of lysine residues
  • UV light-activatable aryl azide group – for crosslinking nonspecifically to the protein side chains and backbone of the interacting protein after allowing protein binding to occur
  • Cleavable disulfide bond (S-S) – can be reduced to release the crosslinker from the original "bait" protein
  • Biotin group – remains attached to target interacting protein after cleaving the disulfide bond, thereby tagging the previously unknown interacting protein(s) for affinity purification and detection

Typical protocol for label transfer experiment:

  1. Add a few microliters of dissolved Sulfo-SBED Reagent to 0.5-1mL of purified bait protein in PBS.
  2. Incubate mixture for 30-120 minutes on ice or at room temperature in the dark.
  3. Desalt or dialyze (in subdued light) to remove excess non-reacted Sulfo-SBED from the labeled bait protein.
  4. Add labeled bait protein to cell lysate or other solution containing putative target protein interactors ("prey").
  5. When interaction complexes have formed, expose the solution to ultraviolet light (365 nm) for several minutes.
  6. Analyze products by one of several methods:
    • Western Blotting: Cleave crosslinks in DTT, separate proteins by SDS-PAGE, and detect biotinylated bands by Western blotting with streptavidin-HRP.
    • Purification and Mass Spec or Sequencing: Affinity-purify biotinylated proteins or peptide fragments following trypsin digestion and perform MS or sequencing to characterize the proteins involved.
Experimental strategy for Sulfo-SBED biotin label transfer and analysis by Western blotting.

References:

  1. Emara M. et al. (2012) Retagging identifies dendritic cell-specific intercellular adhesion molecule-3 (ICAM3)-grabbing non-integrin (DC-SIGN) protein as a novel receptor for a major allergen from house dust mite. J Biol Chem. 287, 5756-5763. PMID: 22205703
  2. Loers G. et al. (2012) The interaction between cell adhesion molecule L1, matrix metalloproteinase 14, and adenine nucleotide translocator at the plasma membrane regulates L1-mediated neurite outgrowth of murine cerebellar neurons. J Neurosci. 32, 3917-3930.PMID: 22423112
  3. Moore B. et al. (2011) Structural basis of molecular recognition of the Leishmania small hydrophilic endoplasmic reticulum-associated protein (SHERP) at membrane surfaces. J Biol Chem. 286, 9246-9256.PMID: 21106528
  4. Ishmael, F.T., et al. (2003). Protein-protein interactions in the bacteriophage T4 replisome. The leading strand holoenzyme is physically linked to the lagging strand holoenzyme and the primosome. J. Biol. Chem. 278: 3145-3152.
  5. Bower, K., et al. (2003). Cell surface antigens of Mycoplasma species bovine group 7 bind to and activate plasminogen. Infect. Immuno. 71: 4823-4827.
  6. Ilver, D., et al. (2003). Bacterium-host protein-carbohydrate interactions. Methods Enzymol. 363: 134-157.
  7. Neely, K.E., et al. (2002). Transcription activator interactions with multiple SWI/SNF subunits. Mol. Cell. Biol., 22(6): 1615-1625.
  8. Tubbs, C.E., et al. (2002). Binding of protein D/E to the surface of rat epidiymal sperm before ejaculation and after deposition in the female reproductive tract. J. Andrology, 23(4): 512-521.
  9. Ishmael, F.T., et al. (2002). Assembly of the bacteriophage T4 helicase-architecture and stoichiometry of the gp41-gp59 complex. J. Biol. Chem. 277: 20555-20562.
  10. Santhoshkumar, P. and Sharma, K.K. (2002). Identification of a region in alcohol dehydrogenase that binds to α-crystallin during chaperone action. Biochemica et Biophysica Acta 1589: 115-121.
  11. Muroi, M., et al. (2002). Regions of the mouse CD14 molecule required for toll-like receptor 2-and 4-mediated activation of NF-κB. J. Biol. Chem. 277: 42372-42379.
  12. Yurchenko, V., et al. (2002). Active site residues of cyclophilin A are crucial for its signaling activity via CD147. J. Biol. Chem. 277: 22959-22965.
  13. Alley, S.C., et al. (2000). Mapping protein-protein interactions in the bacteriophage T4 DNA polymerase holoenzyme using a novel trifunctional photo-crosslinking and affinity reagent. J. Am. Chem. Soc. 122: 6126-6127.
  14. Trotman, L.C., et al. (2001). Import of adenovirus DNA involves the nuclear pole complex receptor CAN/Nup214 and histone H1. Nature Cell Biology 3(Dec.): 1092-1100.
  15. Horney, M.J., et al. (2001). Synthesis and characterization of insulin-like growth factor (IGF)-1 photoprobes selective for the IgG-binding proteins (IGFBPs); photoaffinity labeling of the IGF-binding domain on IGFBP-2. J. Biol. Chem. 276(4): 2880-2889.
  16. Daum, J.R., et al. (2000). The 3F3/2 anti-phosphoepitope antibody binds the mitotically phosphorylated anaphase-promoting complex/cyclosome. Current Biology 10(23): R850-857, S1-S2.
  17. Kleene, R., et al. (2000). SH3 binding sites of ZG29p mediate an interaction with amylase and are involved in condensation sorting in the exocrine rat pancreas. Biochemistry 39: 9893-9900.
  18. Minami, Y., et al. (2000). A critical role for the proteasome activator PA28 in the Hsp90-dependent protein refolding. J. Biol. Chem. 275(12): 9055-9061.
  19. Sharma, K.K., et al. (2000). Synthesis and characterization of a peptide identified as a functional element in aA-crystallin. J. Biol. Chem. 275(6): 3767-3771.
  20. Ilver, D., et al. (1998). Helicobactor pylori adhesin binding fucosylated histo-blood group antigens revealed by re-tagging. Science 279(5349): 373-377.
  21. Jacobson, K.A., et al. (1995). Molecular probes for muscarinic receptors: functionalized congeners of selective muscarinic antagonists. Life Sciences 56(11/12): 823-830.
  22. Geselowitz, D.A. and Neumann, R.D. (1995). Quantitation of triple-helix formation using a photo-crosslinkable aryl azide/biotin/oligonucleotide conjugate. BioConjuate Chem. 6: 502-506.

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Ordering Information
 
Product # Description Pkg. Size Instructions MSDS CofA Price
33073 Sulfo-SBED Biotin Label Transfer Kit - Western Blot Application
Formulation: Multi-component kit with Sulfo-SBED reagent
Sufficient For: 8 label transfer reactions for subsequent Western blot analysis

Kit Contents:
Sulfo-SBED, 8 x 1mg
PBS pack (makes 500mL), 1 pack
Label Transfer Buffer (20X), 200mL
Streptavidin-HRP Conjugate, 0.1mg
Dithiothreitol (DTT), 8 x 7.7mg
Slide-A-Lyzer MINI Dialysis Devices Plus Float, 10K MWCO, 10-device kit
8-rxn kit Product Instructions for product #33073 Sulfo-SBED Biotin Label Transfer Kit - Western Blot Application MSDS for product #33073 Sulfo-SBED Biotin Label Transfer Kit - Western Blot Application Certificate of Analysis for product #33073 Sulfo-SBED Biotin Label Transfer Kit - Western Blot Application $497.00

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33033 Sulfo SBED Biotin Label Transfer Reagent
Formulation: Sulfo-N-hydroxysuccinimidyl-2-(6-[biotinamido]-2-(p-azido benzamido)-hexanoamido) ethyl-1,3'-dithioproprionate; powder solid
Sufficient For: 23mL of typical 1X (0.5mM) reaction solution, or approx. 5 uses at 2mg reagent per use
10mg Product Instructions for product #33033 Sulfo SBED Biotin Label Transfer Reagent MSDS for product #33033 Sulfo SBED Biotin Label Transfer Reagent Certificate of Analysis for product #33033 Sulfo SBED Biotin Label Transfer Reagent $257.00

Add to cart
33034 Sulfo-SBED Biotin Label Transfer Reagent, No-Weigh Format
Formulation: Sulfo-N-hydroxysuccinimidyl-2-(6-[biotinamido]-2-(p-azido benzamido)-hexanoamido) ethyl-1,3'-dithioproprionate; powder solid
Sufficient For: 18mL of typical 1X (0.5mM) reaction solution (total), or 8 separate uses
8 x 1mg Product Instructions for product #33034 Sulfo-SBED Biotin Label Transfer Reagent, No-Weigh Format MSDS for product #33034 Sulfo-SBED Biotin Label Transfer Reagent, No-Weigh Format Certificate of Analysis for product #33034 Sulfo-SBED Biotin Label Transfer Reagent, No-Weigh Format $317.00

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Instructions | MSDS | CofA
Product Instructions | MSDS | CofA  

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