NMR studies revealed structural basis for the inhibitory effects of ubistatins in the ubiquitin-mediated signaling pathways - CNRS - Centre national de la recherche scientifique Accéder directement au contenu
Communication Dans Un Congrès Année : 2018

NMR studies revealed structural basis for the inhibitory effects of ubistatins in the ubiquitin-mediated signaling pathways

Résumé

Ubiquitination is a critical protein post-translational modification involved in a variety of vital processes in eukaryotic cells. The discovery of ubistatins [1], small molecules that impair proteasomal degradation of proteins by directly binding to (poly)ubiquitin upstream of the proteasome, makes ubiquitin itself a potential therapeutic target. Although ubistatins have the potential for drug development and clinical applications, the lack of structural details of ubiquitin-ubistatin interactions has impeded their development. To address this deficiency, a panel of new ubistatin derivatives was synthesized and characterized using functional and NMR-based binding assays [2]. We found that the most active compounds contain strongly acidic groups. We then used NMR and small-angle neutron scattering (SANS) to determine the structures of ubiquitin complexes with ubistatin B and hemi-ubistatin B. These structures revealed direct interactions of ubistatins with ubiquitin’s hydrophobic surface-patch and the basic/polar residues surrounding it, which were confirmed by site-directed mutagenesis. Our results show that ubistatin B binds ubiquitin and di-ubiquitin tighter than a high-affinity ubiquitin-receptor, the UBA domain from the proteasomal shuttle protein ubiquilin-1, and shows clear preference for ubiquitin chains linked via K48 over those linked via K11 or K63. The 15N relaxation and SANS data revealed unexpected binding stoichiometries and structural arrangements of ubiquitin or di-ubiquitins in those complexes. Furthermore, through binding to ubiquitin, ubistatin B shields ubiquitin conjugates from disassembly by a range of deubiquitinases, including the 26S proteasome. Finally, we found that ubistatin B penetrates human cancer cells and perturbs the cellular ubiquitin landscape. These findings highlight versatile properties of ubistatins and have implications for their future development and use in targeting ubiquitin-mediated signaling pathways. Combined with the earlier observations that ubistatins can arrest the cell cycle, producing effects similar to proteasome inhibitors [1]), our structural data suggest that the ubiquitin signal is a plausible candidate for therapeutic intervention in the ubiquitin-proteasome pathway.

Domaines

Chimie
Fichier non déposé

Dates et versions

hal-01862632 , version 1 (27-08-2018)

Identifiants

  • HAL Id : hal-01862632 , version 1

Citer

Mark Nakasone, Thimothy Lewis, Olivier Walker, Jennifer L. Goeckeler-Fried, Daoning Zhang, et al.. NMR studies revealed structural basis for the inhibitory effects of ubistatins in the ubiquitin-mediated signaling pathways. XXVIII International Conference on Magnetic Resonance in Biological Systems (ICMRBS), University College Dublin, Aug 2018, Dublin, Ireland. ⟨hal-01862632⟩
78 Consultations
0 Téléchargements

Partager

Gmail Facebook X LinkedIn More