• My NEB
  • Print
  • PDF
    •

    CoA-SH

    Description

    CoA-SH is a reactive building block for the one-step synthesis of ACP-tag or MCP-tag substrates from maleimide-containing precursors including fluorophores, peptides or oligonucleotides. It is suitable for linkage to maleimides and iodoacetamides.

    The ACP-tag and MCP-tag are polypeptide tags (8 kD) based on the acyl carrier protein. MCP-tag contains two mutations (D36T and D39G). Both allow the specific, covalent attachment of virtually any molecule to a protein of interest. Substrates are derivates of Coenzyme A (CoA). In the labeling reaction, the substituted phosphopantetheine group of CoA is covalently attached to a conserved serine residue of the ACP-tag or the MCP-tag by a phosphopantetheine transferase (SFP Synthase or ACP Synthase).

    While the ACP Synthase (NEB #P9301) will label predominantly the ACP-tag, the SFP Synthase (NEB #P9302) will modify ACP-tag and MCP-tag. CoA substrates react with both the ACP-tag and MCP-tag.

    Having no cysteines, the ACP-tag and the MCP-tag are particularly suited for specifically labeling cell-surface proteins, and should be useful for labeling secreted proteins with disulfide bridges such as antibodies.

    There are two steps to using this system: sub-cloning and expression of the protein of interest as an ACP-tag or MCP-tag fusion, and labeling of the fusion protein using the appropriate synthase with the CoA substrate of choice. In this document, the labeling of the fusion proteins with CoA substrates is described. The cloning of ACP-tag and MCP-tag protein fusions is described in the documentation supplied with the ACP-tag or MCP-tag plasmids.

    Figure 1. Structure of CoA-SH (MW 785.3 g/mol)

    Properties and Usage

    Storage Temperature

    -20°C

    Quality Control

    Quality Control Assays

    The following Quality Control Tests are performed on each new lot and meet the specifications designated for the product. Individual lot data can be found on the Product Summary Sheet/Datacard or Manual which can be found in the Supporting Documents section of this page.
    • Physical Purity (HPLC):
      The purity of the product is determined by HPLC analysis.

    Notes

    1. Storage:
      Store the vials containing the CoA substrate at -20°C. The CoA-SH building block should be dissolved in Tris buffer pH 7.5. Choose a solvent volume that will be compatible with the final use. Unused building block in Tris buffer pH 7.5 should be stored at -20°C under argon.

    References

    1. Yin, J. et al. (2005). Chemistry & Biology. 12, 999-1006.
    2. George, N. et al. (2004). JACS. 126, 8896-8897.
    3. Vivero-Pol, L. et al. (2005). JACS. 127, 12770-12771.

    Supporting Documents

    Material Safety Datasheets

    The following is a list of Material Safety Data Sheets (MSDS) that apply to this product to help you use it safely. The following file naming structure is used to name these document files: [Product Name] MSDS. For international versions please contact us at info@neb.com.
    1. Cellular Imaging and Analysis FAQs
    1. Reaction Conditions for Chemical Coupling with CoA-SH (S9352S)
    2. Protocol for Labeling ACP- or MCP-tag Fusion Proteins with COA-SH Substrates (S9352)

    Selection Tools

    Troubleshooting Guides