Abstract
Function of intrinsically disordered proteins may depend on deviation of their conformational ensemble from that of a random coil. Such deviation may be hard to characterize and quantify, if it is weak. We explored the potential of distance distributions between spin labels, as they can be measured by electron paramagnetic resonance techniques, for aiding such characterization. On the example of the intrinsically disordered N-terminal domain 1–267 of fused in sarcoma (FUS) we examined what such distance distributions can and cannot reveal on the random-coil reference state. On the example of the glycine-rich domain 188–320 of heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1) we studied whether deviation from a random-coil ensemble can be robustly detected with 19 distance distribution restraints. We discuss limitations imposed by ill-posedness of the conversion of primary data to distance distributions and propose overlap of distance distributions as a fit criterion that can tackle this problem. For testing consistency and size sufficiency of the restraint set, we propose jack-knife resampling. At current desktop computers, our approach is expected to be viable for domains up to 150 residues and for between 10 and 50 distance distribution restraints.
Highlights
Based on Anfinsen’s influential thermodynamic hypothesis, function of proteins was considered for a long time to depend exclusively on domains that are well represented by a single conformer with minimum free energy
Probing the Reference State by double electron resonance (DEER) Distance Distributions Before we can address the problem of weak structure, we need to establish what can be inferred by DEER distance distributions on an unstructured protein
We analyze this on the example of the N-terminal domain (NTD) of fused in sarcoma (FUS), where we used protein constructs that do not contain any well-structured domains and are well suited as a fully disordered reference system
Summary
Based on Anfinsen’s influential thermodynamic hypothesis, function of proteins was considered for a long time to depend exclusively on domains that are well represented by a single conformer with minimum free energy. Structure of such domains can be specified at atomic resolution and can be characterized by well-established techniques, such as x-ray crystallography, NMR spectroscopy, and cryo-electron microscopy. IDDs and IDPs are described as structure-less, implying that they exhibit random-coil behavior similar to the one postulated for chemically denatured proteins (Fitzkee and Rose, 2004) This assumption is at odds with the observation that many disordered domains contain evolutionary conserved residues and that mutations in such domains can be pathogenic. This applies even to disordered domains that have not been found to fold upon binding to other proteins or nucleic acids, suggesting that conformation space of proteins cannot be described by a dichotomy of atomic-resolution structure and complete
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