Cell-type specific covalent modifications of proteins in vivo

This technology is an in vivo system in which a covalent bond can be formed between any two proteins in a cell-type specific manner for the purpose of immunoprecipitation, visualization, or modifying the activity of a target protein.

Unmet Need: Simple in vivo tagging technique to covalently link two proteins

Current methods to fuse proteins to other proteins or DNA for immunoprecipitation or visualization are not cell-type specific. Whole tissue chromatin immunoprecipitation (ChIP), for example, is not able to characterize protein-DNA binding in specific cell types within complex tissues. Using a tagging system to covalently bind two proteins of interest in vivo is also not cell-type specific and may require large antibodies. There are currently no assay platforms available that enable direct covalent bonding of two peptides in vivo with cell type-specificity without interfering with native protein structure, localization, or function.

The Technology: Cell-type specific, adaptable assay for covalently linking proteins in vivo

This assay is based on the already existing SpyTag-SpyCatcher system whereby a peptide tag forms a rapid covalent bond to a protein, through engineering a bacterial adhesin. The effector protein can be an epitope, fluorescent protein, or enzyme. The “SpyCatcher::effector” protein is only expressed in the cell type of interest. By co-expressing the SpyTagged protein and the SpyCatcher-tagged effector in a specific cell type, the two tags form a covalent bond in vivo, fusing the two proteins.

This technology has been validated in Drosophila melanogaster tissues.

Applications:

  • Research tool for studying protein function and interactions
  • Tool for manipulating the long-range interactions between proteins
  • Functionalization protocol for lab on chip, enzyme cyclization, or personalized therapeutics
  • Antigen delivery for vaccines
  • Induced degradation tag
  • Epitope tagging
  • Adding functional groups to proteins

Advantages:

  • Cost-effective
  • Cell type-specific
  • Can be used in many types of organisms and tissues
  • Doesn’t require tissue dissociation or nuclei sorting
  • Covalent bonds are stronger than non-covalent protein-protein or protein-small molecule interactions

Lead Inventor:

Richard Mann, Ph.D.

Related Publications:

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