Dataset Information

Conformation and Dynamics of the Troponin I C-Terminal Domain: Combining Single-Molecule and Computational Approaches for a Disordered Protein Region.

ABSTRACT: In recent years, single-molecule Förster resonance energy transfer (smFRET) has emerged as a critical and flexible tool in structural biology, particularly in the study of highly dynamic regions and molecular assemblies. The usefulness of smFRET can be further extended by combining it with computational approaches, marrying the coarse-grained experimental data with higher-resolution in silico calculations. Here we use smFRET to determine six pairwise distances within the intrinsically disordered C-terminal domain of the troponin I subunit (TnIC) of the cardiac troponin complex. We used published conflicting structures of TnIC as starting models for molecular dynamics simulations, which were validated through successful comparison with smFRET measurements before extracting information on conformational dynamics. We find that pairwise distances between residues fluctuate widely in silico, but simulations are generally in good agreement with longer time scale smFRET measurements after averaging across time. Finally, Monte Carlo simulations establish that the lower-energy conformers of TnIC are indeed varied, but that the highest-sampled clusters resemble the published, conflicting models. In this way, we find that the controversial structures are simply stabilized local minima of this dynamic region, and a population including all three would still be consistent with spectroscopic measurements. Taken together, the combined approaches described here allow us to critically evaluate existing models of TnIC, giving insight into the conformation and dynamics of TnIC's disordered state prior to its probable disorder-order transition. Moreover, they provide a framework for combining computational and experimental methods with different time scales for the study of disordered and dynamic protein states.

SUBMITTER: Metskas LA

PROVIDER: S-EPMC4697935 | biostudies-literature | 2015 Sep

REPOSITORIES: biostudies-literature

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Publications

Conformation and Dynamics of the Troponin I C-Terminal Domain: Combining Single-Molecule and Computational Approaches for a Disordered Protein Region.

Metskas Lauren Ann LA Rhoades Elizabeth E

Journal of the American Chemical Society 20150910 37

In recent years, single-molecule Förster resonance energy transfer (smFRET) has emerged as a critical and flexible tool in structural biology, particularly in the study of highly dynamic regions and molecular assemblies. The usefulness of smFRET can be further extended by combining it with computational approaches, marrying the coarse-grained experimental data with higher-resolution in silico calculations. Here we use smFRET to determine six pairwise distances within the intrinsically disordered ...[more]

PMID: 26327565

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Project description:Intrinsically disordered proteins (IDPs) or intrinsically disordered regions (IDRs) is a class of biologically important proteins exhibiting specific biophysical characteristics. They lack a hydrophobic core, and their conformational behavior is strongly influenced by electrostatic interactions. IDPs and IDRs are highly dynamic, and a characterization of the motions of IDPs and IDRs is essential for their physically correct description. NMR together with molecular dynamics simulations are the methods best suited to such a task because they provide information about dynamics of proteins with atomistic resolution. Here, we present a study of motions of a disordered C-terminal domain of the delta subunit of RNA polymerase from Bacillus subtilis. Positively and negatively charged residues in the studied domain form transient electrostatic contacts critical for the biological function. Our study is focused on investigation of ps-ns dynamics of backbone of the delta subunit based on analysis of amide 15N NMR relaxation data and molecular dynamics simulations. In order to extend an informational content of NMR data to lower frequencies, which are more sensitive to slower motions, we combined standard (high-field) NMR relaxation experiments with high-resolution relaxometry. Altogether, we collected data reporting the relaxation at 12 different magnetic fields, resulting in an unprecedented data set. Our results document that the analysis of such data provides a consistent description of dynamics and confirms the validity of so far used protocols of the analysis of dynamics of IDPs also for a partially folded protein. In addition, the potential to access detailed description of motions at the timescale of tens of ns with the help of relaxometry data is discussed. Interestingly, in our case, it appears to be mostly relevant for a region involved in the formation of temporary contacts within the disordered region, which was previously proven to be biologically important.

Dataset Information

Conformation and Dynamics of the Troponin I C-Terminal Domain: Combining Single-Molecule and Computational Approaches for a Disordered Protein Region.

Publications

Conformation and Dynamics of the Troponin I C-Terminal Domain: Combining Single-Molecule and Computational Approaches for a Disordered Protein Region.

Similar Datasets

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