Experimental And Computational Analysis Of The Structure And Dynamics Of Intrinsically Disordered Proteins

Intrinsically disordered proteins (IDPs) are abundant in cells and have central roles in protein-protein interaction networks. Many are involved in cancer, aging and neurodegenerative diseases. The structure and dynamics of IDPs is intimately related to their interactions with binding partners. Because IDPs are inherently flexible and do not have a single conformation, conventional methods and conditions for determining structure and dynamics of globular proteins may not be directly applicable. Nuclear magnetic resonance (NMR) spectroscopy is one of the primary techniques characterizing the structures and dynamics of IDPs, but one cannot rely solely on NMR data. A primary aim of this work was to use Molecular Dynamics (MD) simulations in conjunction with NMR and other biophysical techniques to achieve a deeper understanding of the structure and dynamics of IDPs. To establish suitable parameters and force field choice for simulating IDPs, extensive MD simulations were performed and the results were compared to experimental data. Using computational and experimental techniques, the interactions between peptides from 9 disordered proteins with a common target were interrogated. The findings allowed us to determine key factors in modulating the affinities of the various interactions and highlighted the importance of molecular recognition fragments (MoRFs) in IDP target recognition and binding. IDP binding was also investigated from the perspective of the binding partner. The backbone resonances of the 3̃2 kDa target were assigned and the binding interface was mapped in the presence of a peptide from a disordered binding partner. Chemical shift changes distant from the interaction site indicated that IDP binding is a complex process, which should be studied from the perspectives of the partner and target. Because IDPs are highly sensitive to environmental conditions, the effects of molecular crowding on the dynamics of IDPs were also investigated. I found that crowding might have differential effects on the conformational propensities of distinct regions of some IDPs. This information will help to understand the behavior of IDPs in cellular environments and to determine suitable conditions for accurately studying them. This work has helped to improve the understanding of how IDP structure and dynamics relate to target binding.