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Characterization of Three Druggable Hot-Spots in the Aurora-A/TPX2 Interaction Using Biochemical, Biophysical, and Fragment-Based Approaches.

McIntyre PJ, Collins PM, Vrzal L, Birchall K, Arnold LH, Mpamhanga C, Coombs PJ, Burgess SG, Richards MW, Winter A, Veverka V, Delft FV, Merritt A, Bayliss R
ACS Chem Biol 12 2906-2914 (2017)
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Cited: 21 times
EuropePMC logo PMID: 29045126

Abstract

The mitotic kinase Aurora-A and its partner protein TPX2 (Targeting Protein for Xenopus kinesin-like protein 2) are overexpressed in cancers, and it has been proposed that they work together as an oncogenic holoenzyme. TPX2 is responsible for activating Aurora-A during mitosis, ensuring proper cell division. Disruption of the interface with TPX2 is therefore a potential target for novel anticancer drugs that exploit the increased sensitivity of cancer cells to mitotic stress. Here, we investigate the interface using coprecipitation assays and isothermal titration calorimetry to quantify the energetic contribution of individual residues of TPX2. Residues Tyr8, Tyr10, Phe16, and Trp34 of TPX2 are shown to be crucial for robust complex formation, suggesting that the interaction could be abrogated through blocking any of the three pockets on Aurora-A that complement these residues. Phosphorylation of Aurora-A on Thr288 is also necessary for high-affinity binding, and here we identify arginine residues that communicate the phosphorylation of Thr288 to the TPX2 binding site. With these findings in mind, we conducted a high-throughput X-ray crystallography-based screen of 1255 fragments against Aurora-A and identified 59 hits. Over three-quarters of these hits bound to the pockets described above, both validating our identification of hotspots and demonstrating the druggability of this protein-protein interaction. Our study exemplifies the potential of high-throughput crystallography facilities such as XChem to aid drug discovery. These results will accelerate the development of chemical inhibitors of the Aurora-A/TPX2 interaction.

Articles - 5oro mentioned but not cited (1)

  1. research-article AlphaFold2 models of the active form of all 437 catalytically competent human protein kinase domains. Faezov B, Dunbrack RL. bioRxiv 2023.07.21.550125 (2023)


Reviews citing this publication (5)

  1. The multifaceted allosteric regulation of Aurora kinase A. Levinson NM. Biochem. J. 475 2025-2042 (2018)
  2. Protein X-ray Crystallography and Drug Discovery. Maveyraud L, Mourey L. Molecules 25 (2020)
  3. Aurora A and AKT Kinase Signaling Associated with Primary Cilia. Nishimura Y, Yamakawa D, Shiromizu T, Inagaki M. Cells 10 3602 (2021)
  4. Aurora Kinase A Regulation by Cysteine Oxidative Modification. Lee IG, Lee BJ. Antioxidants (Basel) 12 531 (2023)
  5. Structural features of the protein kinase domain and targeted binding by small-molecule inhibitors. Arter C, Trask L, Ward S, Yeoh S, Bayliss R. J Biol Chem 298 102247 (2022)

Articles citing this publication (15)

  1. A dynamic mechanism for allosteric activation of Aurora kinase A by activation loop phosphorylation. Ruff EF, Muretta JM, Thompson AR, Lake EW, Cyphers S, Albanese SK, Hanson SM, Behr JM, Thomas DD, Chodera JD, Levinson NM. Elife 7 (2018)
  2. Bora phosphorylation substitutes in trans for T-loop phosphorylation in Aurora A to promote mitotic entry. Tavernier N, Thomas Y, Vigneron S, Maisonneuve P, Orlicky S, Mader P, Regmi SG, Van Hove L, Levinson NM, Gasmi-Seabrook G, Joly N, Poteau M, Velez-Aguilera G, Gavet O, Castro A, Dasso M, Lorca T, Sicheri F, Pintard L. Nat Commun 12 1899 (2021)
  3. Construction of a Shape-Diverse Fragment Set: Design, Synthesis and Screen against Aurora-A Kinase. Zhang R, McIntyre PJ, Collins PM, Foley DJ, Arter C, von Delft F, Bayliss R, Warriner S, Nelson A. Chemistry 25 6831-6839 (2019)
  4. Dynamics of human protein kinase Aurora A linked to drug selectivity. Pitsawong W, Buosi V, Otten R, Agafonov RV, Zorba A, Kern N, Kutter S, Kern G, Pádua RA, Meniche X, Kern D. Elife 7 (2018)
  5. A Thermodynamic Model for Multivalency in 14-3-3 Protein-Protein Interactions. Stevers LM, de Vink PJ, Ottmann C, Huskens J, Brunsveld L. J. Am. Chem. Soc. 140 14498-14510 (2018)
  6. Demonstration of the utility of DOS-derived fragment libraries for rapid hit derivatisation in a multidirectional fashion. Kidd SL, Fowler E, Reinhardt T, Compton T, Mateu N, Newman H, Bellini D, Talon R, McLoughlin J, Krojer T, Aimon A, Bradley A, Fairhead M, Brear P, Díaz-Sáez L, McAuley K, Sore HF, Madin A, O'Donovan DH, Huber KVM, Hyvönen M, von Delft F, Dowson CG, Spring DR. Chem Sci 11 10792-10801 (2020)
  7. High-Throughput Crystallography Reveals Boron-Containing Inhibitors of a Penicillin-Binding Protein with Di- and Tricovalent Binding Modes. Newman H, Krajnc A, Bellini D, Eyermann CJ, Boyle GA, Paterson NG, McAuley KE, Lesniak R, Gangar M, von Delft F, Brem J, Chibale K, Schofield CJ, Dowson CG. J Med Chem 64 11379-11394 (2021)
  8. A new soaking procedure for X-ray crystallographic structural determination of protein-peptide complexes. Ballone A, Lau RA, Zweipfenning FPA, Ottmann C. Acta Crystallogr F Struct Biol Commun 76 501-507 (2020)
  9. Allosteric modulation of a human protein kinase with monobodies. Zorba A, Nguyen V, Koide A, Hoemberger M, Zheng Y, Kutter S, Kim C, Koide S, Kern D. Proc. Natl. Acad. Sci. U.S.A. 116 13937-13942 (2019)
  10. Computational Analysis of Crystallization Additives for the Identification of New Allosteric Sites. Fogha J, Diharce J, Obled A, Aci-Sèche S, Bonnet P. ACS Omega 5 2114-2122 (2020)
  11. Covalent Aurora A regulation by the metabolic integrator coenzyme A. Tsuchiya Y, Byrne DP, Burgess SG, Bormann J, Baković J, Huang Y, Zhyvoloup A, Yu BYK, Peak-Chew S, Tran T, Bellany F, Tabor AB, Chan AE, Guruprasad L, Garifulin O, Filonenko V, Vonderach M, Ferries S, Eyers CE, Carroll J, Skehel M, Bayliss R, Eyers PA, Gout I. Redox Biol 28 101318 (2020)
  12. Fragment-Based Ligand Discovery Applied to the Mycolic Acid Methyltransferase Hma (MmaA4) from Mycobacterium tuberculosis: A Crystallographic and Molecular Modelling Study. Galy R, Ballereau S, Génisson Y, Mourey L, Plaquevent JC, Maveyraud L. Pharmaceuticals (Basel) 14 1282 (2021)
  13. Ligand discrimination between active and inactive activation loop conformations of Aurora-A kinase is unmodified by phosphorylation. Gilburt JAH, Girvan P, Blagg J, Ying L, Dodson CA. Chem Sci 10 4069-4076 (2019)
  14. Novel Aurora A Kinase Inhibitor Fangchinoline Enhances Cisplatin-DNA Adducts and Cisplatin Therapeutic Efficacy in OVCAR-3 Ovarian Cancer Cells-Derived Xenograft Model. Winardi D, Chu PY, Chen GY, Wang K, Hsu WY, Hsieh CL, Chen YH, Wu YC, Yang JC. Int J Mol Sci 23 1868 (2022)
  15. Structural basis for CEP192-mediated regulation of centrosomal AURKA. Park JG, Jeon H, Shin S, Song C, Lee H, Kim NK, Kim EE, Hwang KY, Lee BJ, Lee IG. Sci Adv 9 eadf8582 (2023)


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