title: Model of the Interaction between the NF-κB Inhibitory Protein p100 and the E3 Ubiquitin Ligase β-TrCP based on NMR and Docking Experiments.
abstract: NF-κB is a major transcription factor whose activation is triggered through two main activation pathways: the canonical pathway involving disruption of IκB-α/NF-κB complexes and the alternative pathway whose activation relies on the inducible proteolysis of the inhibitory protein p100. One central step controlling p100 processing consists in the interaction of the E3 ubiquitin ligase β-TrCP with p100, thereby leading to its ubiquitinylation and subsequent either complete degradation or partial proteolysis by the proteasome. However, the interaction mechanism between p100 and β-TrCP is still poorly defined. In this work, a diphosphorylated 21-mer p100 peptide model containing the phosphodegron motif was used to characterize the interaction with β-TrCP by NMR. In parallel, docking simulations were performed in order to obtain a model of the 21P-p100/β-TrCP complex. Saturation transfer difference (STD) experiments were performed in order to highlight the residues of p100 involved in the interaction with the β-TrCP protein. These results highlighted the importance of pSer865and pSer869residues in the interaction with β-TrCP and particularly the Tyr867that fits inside the hydrophobe β-TrCP cavity with the Arg474 guanidinium group. Four other arginines, Arg285, Arg410, Arg431, and Arg521, were found essential in the stabilization of p100 on the β-TrCP surface. Importantly, the requirement for these five arginine residues of β-TrCP for the interaction with p100 was further confirmed in vivo, thereby validating the docking model through a biological approach.
authors.affiliation: Laboratoire de Chimie et Biochimie Pharmacologiques et Toxicologiques, UMR 8601-CNRS, Université Paris Descartes , Sorbonne Paris Cité, 45 rue des Saint-Pères, 75006, Paris, France.
authors.forename: Maxime
authors.initials: M
authors.lastname: Melikian
authors.affiliation: NF-kB, Differentiation and Cancer, Université Paris Descartes , Sorbonne Paris Cité, 4 Avenue de l'Observatoire, 75006, Paris, France.
authors.forename: Baptiste
authors.initials: B
authors.lastname: Eluard
authors.affiliation: Laboratoire de Chimie et Biochimie Pharmacologiques et Toxicologiques, UMR 8601-CNRS, Université Paris Descartes , Sorbonne Paris Cité, 45 rue des Saint-Pères, 75006, Paris, France.
authors.forename: Gildas
authors.initials: G
authors.lastname: Bertho
authors.affiliation: NF-kB, Differentiation and Cancer, Université Paris Descartes , Sorbonne Paris Cité, 4 Avenue de l'Observatoire, 75006, Paris, France.
authors.forename: Véronique
authors.initials: V
authors.lastname: Baud
authors.affiliation: Laboratoire de Chimie et Biochimie Pharmacologiques et Toxicologiques, UMR 8601-CNRS, Université Paris Descartes , Sorbonne Paris Cité, 45 rue des Saint-Pères, 75006, Paris, France.
authors.forename: Nathalie
authors.identifier: 0000-0003-1210-5750
authors.initials: N
authors.lastname: Evrard-Todeschi
mesh_terms: D000595:Amino Acid Sequence; D009682:Magnetic Resonance Spectroscopy; D062105:Molecular Docking Simulation; D056004:Molecular Dynamics Simulation; D009154:Mutation; D052003:NF-kappa B p52 Subunit / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D044844:beta-Transducin Repeat-Containing Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D052003:NF-kappa B p52 Subunit; D044844:beta-Transducin Repeat-Containing Proteins