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Fine-tuning probes for fluorescence polarization binding assays of bivalent ligands against polo-like kinase 1 using full-length protein

  1. Author:
    Tsuji, Kohei
    Tamamura, Hirokazu
    Burke,Terrence

  2. Author Address

    Department of Medicinal Chemistry, Laboratory for Biomaterials and Bioengineering, Institute of Integrated Research, Institute of Science Tokyo, 2-3-10 Kandasurugadai, Chiyoda-ku, Tokyo 101-0062, Japan; Chemical Biology Laboratory, Center for Cancer Research, National Cancer Institute, National Institutes of Health, 1050 Boyles St., Frederick, MD 21702, USA. Electronic address: ktsuji.mr@tmd.ac.jp., Department of Medicinal Chemistry, Laboratory for Biomaterials and Bioengineering, Institute of Integrated Research, Institute of Science Tokyo, 2-3-10 Kandasurugadai, Chiyoda-ku, Tokyo 101-0062, Japan.,
    1. Year: 2024
    2. Date: Dec 28
    3. Epub Date: 2024 12 28
  2. Journal: Bioorganic & Medicinal Chemistry
    1. 119
    2. Pages: 118055
  4. Type of Article: Article
  5. Article Number: 118055
  1. Abstract:

    Polo-like kinase 1 (Plk1) is an important cell cycle regulator that is a recognized target for development of anti-cancer therapeutics. Plk1 is composed of a catalytic kinase domain (KD), a flexible interdomain linker and a polo-box domain (PBD). Intramolecular protein-protein interactions (PPIs) between the PBD and KD result in "auto-inhibition" that is an essential component of proper Plk1 function. Recently, we developed high-affinity PBD-binding inhibitors using a bivalent approach. These ligands contain the low-nanomolar affinity Plk1 KD-binding inhibitors BI2536 or Wortmannin tethered to the PBD-binding peptide, PLH*SpT (H* represents a -(CH2)8Ph group on the histidine side chain p-nitrogen). Due to the extremely high affinity of these bivalent inhibitors, to avoid bottoming out in competitive binding assays, it was necessary to use PLH*SpT in the affinity probe. As reported herein, we have developed fluorescence polarization assays using a new fluorescent probe based on the Plk1 PBD-binding peptide, FDPPLHSpTA. We applied the assay to evaluate the affinities of bivalent inhibitors that possess a variety of PBD-binding peptides having much lower PBD-affinities than PLH*SpT. Tethering BI2536 in these bivalent inhibitors resulted in significant affinity enhancements as compared to the parent monovalent peptides. Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.

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External Sources

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  2. DOI: 10.1016/j.bmc.2024.118055
  3. PMID: 39764864
  4. PII : S0968-0896(24)00469-3

Library Notes

  1. Fiscal Year: FY2024-2025
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