Cited by 50NF-κB Signaling PathwaysComputational Drug Discovery MethodsProtein Structure and Dynamics

Comprehensive Experimental and Computational Analysis of Binding Energy Hot Spots at the NF-κB Essential Modulator/IKKβ Protein–Protein Interface

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Golden, Mary S., Cote, Shaun M., Sayeg, Marianna, Zerbe, Brandon S., Villar, Elizabeth A., Beglov, Dmitri, Sazinsky, Stephen L., Georgiadis, Rosina M. +3 more·2013·Journal of the American Chemical Society

Abstract

We report a comprehensive analysis of binding energy hot spots at the protein-protein interaction (PPI) interface between nuclear factor kappa B (NF-κB) essential modulator (NEMO) and IκB kinase subunit β (IKKβ), an interaction that is critical for NF-κB pathway signaling, using experimental alanine scanning mutagenesis and also the FTMap method for computational fragment screening. The experimental results confirm that the previously identified NEMO binding domain (NBD) region of IKKβ contains the highest concentration of hot-spot residues, the strongest of which are W739, W741, and L742 (ΔΔG = 4.3, 3.5, and 3.2 kcal/mol, respectively). The region occupied by these residues defines a potentially druggable binding site on NEMO that extends for ~16 Å to additionally include the regions that bind IKKβ L737 and F734. NBD residues D738 and S740 are also important for binding but do not make direct contact with NEMO, instead likely acting to stabilize the active conformation of surrounding residues. We additionally found two previously unknown hot-spot regions centered on IKKβ residues L708/V709 and L719/I723. The computational approach successfully identified all three hot-spot regions on IKKβ. Moreover, the method was able to accurately quantify the energetic importance of all hot-spot residues involving direct contact with NEMO. Our results provide new information to guide the discovery of small-molecule inhibitors that target the NEMO/IKKβ interaction. They additionally clarify the structural and energetic complementarity between "pocket-forming" and "pocket-occupying" hot-spot residues, and further validate computational fragment mapping as a method for identifying hot spots at PPI interfaces.

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  1. [2]Berg T Curr. Opin. Drug Disc. Develop 2008 11 666 18729018
  2. [44]Feinman R Siegel DS Berenson J Clin. Adv. Hematol. Oncol 2004 2 162 16166946

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Authors
Golden, M. S., Cote, S. M., Sayeg, M., Zerbe, B. S., Villar, E. A., Beglov, D., Sazinsky, S. L., Georgiadis, R. M., Vajda, S., Kozakov, D., & Whitty, A.
Affiliations
  • Boston University (US)
  • Departments of †Chemistry and ‡Biomedical Engineering, Boston University, 590 Commonwealth Avenue, Boston, Massachusetts 02215, United States
Corresponding author
  • Kozakov, Dima
  • Whitty, Adrian
Venue
Journal of the American Chemical Society
Published
2013-04-10
Volume
135
Issue
16
Pages
6242-6256
ISSN
  • 0002-7863 (ISSN-L)
  • 1520-5126
OpenAlex
W2035775380
Cited by
47
References
74
Citations by year
2026: 3 · 2024: 3 · 2023: 1 · 2022: 3 · 2021: 2 · 2020: 3 · 2019: 6 · 2018: 4 · 2017: 2 · 2016: 5 · 2015: 4 · 2014: 8 · 2013: 3
FWCI
2.46
Type
journal article
Language
en
Open access
Locations
  • Journal of the American Chemical Society (publishedVersion)
  • PubMed (publishedVersion)
  • Europe PMC (PubMed Central) (acceptedVersion)
  • PubMed Central (acceptedVersion)
Funders
  • National Institutes of Health (4r01gm061867-15)
  • National Institute of General Medical Sciences (GM094551, R01GM094551, GM93147, GM064700, R01GM093147, GM61687, R01GM064700, R01-GM061867)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Cancer Research › NF-κB Signaling Pathways
  • Physical Sciences › Computer Science › Computational Theory and Mathematics › Computational Drug Discovery Methods
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein Structure and Dynamics
Keywords
NEMO, IKKβ, protein-protein interaction, NF-κB signaling, alanine scanning, small molecule inhibitors, binding free energy
MeSH terms
Alanine, Algorithms, Amino Acids, Genetic Vectors, Models, Molecular, Protein Binding, Recombinant Fusion Proteins, X-Ray Diffraction, Signal Transduction, Mutagenesis, Mutagenesis, Site-Directed, NF-kappa B, Anisotropy, Computational Biology, I-kappa B Kinase

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