Discovery of Protein-Protein Interaction Inhibitors by Integrating Protein Engineering and Chemical Screening Platforms

Timurs Maculins, Javier Garcia-Pardo, Anamarija Skenderovic, Jakob Gebel, Mateusz Putyrski, Andrew Vorobyov, Philipp Busse, Gabor Varga, Maria Kuzikov, Andrea Zaliani, Simin Rahighi, Veronique Schaeffer, Michael J. Parnham, Sachdev S. Sidhu, Andreas Ernst, Volker Dötsch, Masato Akutsu, Ivan Dikic

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    4 Citations (Scopus)

    Abstract

    Protein-protein interactions (PPIs) govern intracellular life, and identification of PPI inhibitors is challenging. Roadblocks in assay development stemming from weak binding affinities of natural PPIs impede progress in this field. We postulated that enhancing binding affinity of natural PPIs via protein engineering will aid assay development and hit discovery. This proof-of-principle study targets PPI between linear ubiquitin chains and NEMO UBAN domain, which activates NF-κB signaling. Using phage display, we generated ubiquitin variants that bind to the functional UBAN epitope with high affinity, act as competitive inhibitors, and structurally maintain the existing PPI interface. When utilized in assay development, variants enable generation of robust cell-based assays for chemical screening. Top compounds identified using this approach directly bind to UBAN and dampen NF-κB signaling. This study illustrates advantages of integrating protein engineering and chemical screening in hit identification, a development that we anticipate will have wide application in drug discovery.
    Original languageUndefined/Unknown
    JournalCell Chemical Biology
    DOIs
    Publication statusPublished - Nov 2020

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