Fluorescent Reporters of Protein/Protein and Peptide/Protein Interactions

Applications

The fluorescent reporter of protein/protein and peptide/protein interactions, Dap(4-DMN) (4-dimethylaminonaphthalimide), can be incorporated into any peptide sequence of interest as either a unique amino acid or as a cysteine modification. This fluorophore is sensitive to environmental polarity and can be used to monitor ions, small molecules, and biological processes such as protein folding, protein-protein interactions, and phosphorylation events, among others. 

Problem Addressed

Many currently existing environmentally-sensitive fluorophores lack sensitivity and/or stability, which result in signal noise and degradation. Inventors have previously introduced another fluorophore, Dap(6-DMN), which has a lower chemical stability and a smaller change in fluorescence due to changes in environmental polarity. The Inventors have shown that Dap(4-DMN) undergoes enhanced fluorescence in nonpolar environments compared to Dap(6-DMN). 

Technology

Dap(4-DMN) undergoes enhanced fluorescence in nonpolar environments, due to differences in electron orbital configurations in the molecule that arise from environmental polarity shifts. It absorbs light at approximately 445 nm, and emits from 492-538 nm. Inventors have shown that it can be synthesized as an individual amino acid, which can then be incorporated into the protein or peptide of interest via solid-phase peptide synthesis or protein semi-synthesis. Dap(4-DMN) may also be incorporated into compounds that can conjugate to cysteine in intact peptides and proteins. Photophysical studies have been carried out to demonstrate the superior photophysical properties of Dap(4-DMN), and chemical stability studies have been carried out to reveal advantageous properties for biomolecule labeling and the detection of protein/protein and peptide/protein interactions.

Advantages

  •  Superior sensitivity compared to current environmentally sensitive fluorophores
  •  Chemical stability
  •  Protein incorporation method already developed