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Probing dynamic conformations of the high-molecular-weight αB-crystallin heat shock protein ensemble by NMR spectroscopy.

Authors
  • 1
  • 1 Department of Molecular Genetics, The University of Toronto, Toronto, Ontario M5S 1A8, Canada. [email protected] , (Canada)
Type
Published Article
Journal
Journal of the American Chemical Society
1520-5126
Publisher
American Chemical Society
Publication Date
Volume
134
Issue
37
Pages
15343–15350
Identifiers
PMID: 22916679
Source
Medline
License
Unknown

Abstract

Solution- and solid-state nuclear magnetic resonance (NMR) spectroscopy are highly complementary techniques for studying supra-molecular structure. Here they are employed for investigating the molecular chaperone αB-crystallin, a polydisperse ensemble of between 10 and 40 identical subunits with an average molecular mass of approximately 600 kDa. An IxI motif in the C-terminal region of each of the subunits is thought to play a critical role in regulating the size distribution of oligomers and in controlling the kinetics of subunit exchange between them. Previously published solid-state NMR and X-ray results are consistent with a bound IxI conformation, while solution NMR studies provide strong support for a highly dynamic state. Here we demonstrate through FROSTY (freezing rotational diffusion of protein solutions at low temperature and high viscosity) MAS (magic angle spinning) NMR that both populations are present at low temperatures (<0 °C), while at higher temperatures only the mobile state is observed. Solution NMR relaxation dispersion experiments performed under physiologically relevant conditions establish that the motif interchanges between flexible (highly populated) and bound (sparsely populated) states. This work emphasizes the importance of using multiple methods in studies of supra-molecules, especially for highly dynamic ensembles where sample conditions can potentially affect the conformational properties observed.

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