Combined Experimental And Computational Approaches To Elucidate The Structures And Dynamics Of Flexible Proteins

The function of protein is closely related to protein flexibility. The interaction between a protein and other molecules often requires the protien to change its conformation. This change may be very small, involving only the rearrangement of some side chains, or it may be very large and involve large-scale conformational fluctuations. The studying of protein flexibility is important for describing protein interactions.In the first chapter of this dissertation, there is an introduction of intrinsically disordered proteins (IDPs) and methods for studying them. Over the past few decades, the roles of IDPs have been increasingly recognized despite a lack of well-folded structure. Numerous experimental methods are available to characterise the structural properties of disordered proteins. In particular, nuclear magnetic resonance (NMR) spectroscopy and small-angle X-ray scattering (SAXS) are the most frequently used methods, which provide quantitative structural information.The experimental data can be used in various computational approaches, including restrained molecular dynamics (MD) simulations and other algorithms that can generate structural ensembles of diverse conformers. And the resulting ensembles provide critical insights into the structure and function of IDPs.In Escherichia coli, the RNA chaperone Hfq is an important post-transcriptional regulator. The Hfq protein consists of a conserved Sm domain and an intrinsically disordered C-terminus. Previous studies demonstrate that the C-terminus is essential for the function of Hfq. However, the detailed structural and dynamic properties of the C-terminus are still poorly understood due to the disordered nature of the C-terminus. In the second chapter, we combine experimental and computational approaches to study the structures and dynamics of the C-terminus in intact Hfq. The results demonstrate that the disordered C-terminus is highly flexible and in transient interaction with the hexameric core. Our results also indicate that some regions in the C-terminus are motional restricted and the presence of transient secondary structures. Finally, we present a structure ensemble of the C-terminus in full length Hfq, which provides a structural basis for understanding the role of the C-terminus.Many proteins consist of at least two domains and they are often connected by flexible linkers, which determine the extent of interdomain motions. Large scale conformational flexibility within a multidomain protein is often important for its biological function. Studies of the multi-domain protein with flexible linkers are complicated, leading to methodological challenges. In the third chapter, we develop a method, which combines amplified collective motions (ACM) simulations and SAXS data, to investigate the structural states of multi-domain proteins in solution.The method was applied to two proteins, which are bacteriophage T4 lysozyme and tandem WW domains of the formin-binding protein 21, respectively. The results suggests that the ACM simulations can sample more extensive conformational space of the proteins than the standard MD simulations. Moreover, the structure ensembles generated by ACM are significantly better in reproducing the SAXS data than those from the MD simulations.