Emil Thomasen:
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Understanding the relationship between the structure and biological function of proteins is important for fundamental biology and to develop new therapeutics and biotechnology. Many proteins consist of multiple folded domains connected by flexible linkers or intrinsically disordered regions and display a high level of structural heterogeneity. Determining the conformational states that such a protein adopts can be challenging, and often requires a combination of computational modeling and biophysical experiments. This PhD thesis presents a series of research projects that focus on the integration of coarse-grained (CG) molecular dynamics (MD) simulations and small-angle X-ray scattering (SAXS) experiments to characterize the conformational ensembles of multidomain proteins.
The thesis begins with an introduction to integrative modeling of conformational ensembles of intrinsically disordered proteins (IDPs) and multidomain proteins, followed by a series of articles that present the research projects undertaken throughout the PhD: The first project is focused on characterizing the structure and dynamics of higher-order oligomers of the ubiquitin ligase adaptor protein SPOP. An integrative model based on SAXS and CG MD simulations reveals that SPOP forms linear oligomers with a rigid, helical backbone structure and flexible substrate binding domains. Additionally, the results are consistent with previous evidence that SPOP self-association follows an isodesmic model. The next project investigates the phase behavior of the RNA-binding protein hnRNPA1. An integrative model based on SAXS and CG MD simulations reveals that there are salt-dependent interactions between the folded domains and the disordered low-complexity domain of hnRNPA1, which correlate with its propensity to phase separate. These results illustrate how folded domains can modulate the phase behavior of disordered regions.
The final project is focused on the widely used CG force field Martini 3. The results show that there are discrepancies between Martini 3 simulations and SAXS data for a diverse set of IDPs and multidomain proteins. Increasing the non-bonded interactions between protein and water by 10% or decreasing the non-bonded interactions within proteins by 12% favors more expanded conformations and results in improved agreement with the experimental data. Together, the results presented in this thesis illustrate the power of MD simulations in interpreting SAXS data on dynamic proteins, and the power of SAXS in improving MD simulation methods.