Collision-Induced Unfolding Reveals Unique Fingerprints for Remote Protein Interaction Sites in the KIX Regulation Domain

The kinase-inducible domain (KIX) of the transcriptional coactivator CBP binds multiple transcriptional regulators through two allosterically connected sites. Establishing a method for observing activator-specific KIX conformations would facilitate the discovery of drug-like molecules that capture s...

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Bibliographic Details
Published in:Journal of the American Society for Mass Spectrometry 2019-01, Vol.30 (1), p.94-102
Main Authors: Rabuck-Gibbons, Jessica N., Lodge, Jean M., Mapp, Anna K., Ruotolo, Brandon T.
Format: Article
Language:English
Subjects:
Analytical Chemistry
Binding Sites
Bioinformatics
Biotechnology
Chemistry
Chemistry and Materials Science
Fingerprints
Focus: Honoring Carol V. Robinson's Election to the National Academy of Sciences: Research Article
Ion Mobility Spectrometry - methods
Mass spectrometry
Membrane Proteins - chemistry
Membrane Proteins - metabolism
Multiprotein Complexes - chemistry
Multiprotein Complexes - metabolism
Organic Chemistry
Peptide Mapping - methods
Peptides
Peptides - chemistry
Peptides - metabolism
Phosphoproteins - chemistry
Phosphoproteins - metabolism
Protein Binding
Protein Conformation
Protein Domains
Protein Unfolding
Proteins
Proteomics
Regulators
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Summary:The kinase-inducible domain (KIX) of the transcriptional coactivator CBP binds multiple transcriptional regulators through two allosterically connected sites. Establishing a method for observing activator-specific KIX conformations would facilitate the discovery of drug-like molecules that capture specific conformations and further elucidate how distinct activator-KIX complexes produce differential transcriptional effects. However, the transient and low to moderate affinity interactions between activators and KIX are difficult to capture using traditional biophysical assays. Here, we describe a collision-induced unfolding-based approach that produces unique fingerprints for peptides bound to each of the two available sites within KIX, as well as a third fingerprint for ternary KIX complexes. Furthermore, we evaluate the analytical utility of unfolding fingerprints for KIX complexes using CIUSuite, and conclude by speculating as to the structural origins of the conformational families created from KIX:peptide complexes following collisional activation. Graphical Abstract ᅟ
ISSN:1044-0305
1879-1123
DOI:10.1007/s13361-018-2043-6