Protein structures and interactions are key to understanding their biological function. The current techniques to probe structures, NMR and x-ray crystallography, have limitations that can be overcome with the use of photoaffinity labeling and photooxidative crosslinking. A family of amino-acid functionalized naphthalene imide (NI) and diimide (NDI) compounds do these photoactivatable techniques in order to improve structural based drug design. NI-Alanine covalently crosslinked lysozyme structures as shown by SDS-PAGE while NI-Tyrosine site specifically targeted the active site of mushroom tyrosinase demonstrated by fluorescence and colorimetric enzyme kinetics. When functionalized these naphthalene imide compounds can be used for photooxidative crosslinking to determine protein complex formations. NDI-Alanine non-specifically labeled bovine serum albumin and was verified by HPLC. The proposed labeling mechanism of these compounds involves generating a biradical upon irradiation that photodecarboxylates, leaving a biradical that can react with proteins by a hydrogen abstraction follwed by radical recombination. Photodecarboxylation was verified with mass spectrometry and fluorescence. Radical traps will be utilized to determine the presence of a biradical. Once the mechanism is determined, these naphthalene diimide compounds can be functionalized to probe the structure of active sites of enzymes.

Photooxidative crosslinking and photoaffinity labeling of proteins with naphthalene imides and diimides

254th National Meeting (2017)

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