title: Quaternary dynamics of αB-crystallin as a direct consequence of localised tertiary fluctuations in the C-terminus.
abstract: The majority of proteins exist in vivo within macromolecular assemblies whose functions are dependent on dynamical processes spanning a wide range of time scales. One such assembly is formed by the molecular chaperone αB-crystallin that exists in a variety of exchanging oligomeric states, centred on a mass of approximately 560 kDa. For many macromolecular assemblies, including αB-crystallin, the inherent dynamics, heterogeneity and high mass contribute to difficulties in quantitative studies. Here, we demonstrate a strategy based on correlating solution-state nuclear magnetic resonance spectroscopy and mass spectrometry data to characterize simultaneously the organization and dynamics of the polydisperse αB-crystallin ensemble. We show that protomeric dimers assemble into oligomers via the binding of extended C-termini, with each monomer donating and receiving one terminus. Moreover, we establish that the C-termini undergo millisecond fluctuations that regulate the interconversion of oligomeric forms. The combined biophysical approach allows construction of an energy profile for a single monomer that completely describes the equilibrium dynamics of the ensemble. It also facilitates an analysis of dynamics spanning the millisecond to hour time scales and secondary to quaternary structural levels, and provides an approach for, obtaining simultaneously detailed structural, thermodynamic and kinetic information on a heterogeneous protein assembly.
authors.affiliation: Departments of Molecular Genetics, Biochemistry and Chemistry, The University of Toronto, Toronto, Ontario, Canada M5S 1A8. ajb204@pound.med.utoronto.ca
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Baldwin
authors.forename: Gillian R
authors.initials: GR
authors.lastname: Hilton
authors.forename: Hadi
authors.initials: H
authors.lastname: Lioe
authors.forename: Claire
authors.initials: C
authors.lastname: Bagnéris
authors.forename: Justin L P
authors.initials: JL
authors.lastname: Benesch
authors.forename: Lewis E
authors.initials: LE
authors.lastname: Kay
mesh_terms: D000465:Algorithms; D004574:Electromagnetic Fields; D004926:Escherichia coli / Q000378:metabolism; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D008958:Models, Molecular; D011487:Protein Conformation; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D021122:Protein Subunits; D013816:Thermodynamics; D038203:alpha-Crystallin B Chain / Q000737:chemistry*
chemical_list: D021122:Protein Subunits; D038203:alpha-Crystallin B Chain