Structural and dynamics studies of intrinsically disordered proteins

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The function of a protein is intimately related to its structure, flexibility, and the ways it interacts with other molecules in cells. Studying of the folding, structure, dynamics, and target recognition of a protein is therefore with fundamental importance in understanding how it functions. Intrinsically disordered proteins (IDPs) are a unique class of proteins that do not adopt a well-defined structure but exist as an ensemble of fast inter-converting conformers under physiological conditions. Importantly, such proteins are highly abundant in our bodies and are involved in important biological functions. The long-term goal of my research program is to seek a complete understanding of the complex structure-function relationship of IDPs. We are particularly interested in IDPs that are involved in signaling pathways. To achieve this goal, we will extensively characterize the structural propensity, dynamic feature, and target-binding mechanism of different IDPs with varying structural properties and under different conditions. By using an array of biophysical techniques including nuclear magnetic resonance spectroscopy, isothermal titration calorimetry, circular dichroism spectropolarimety, as well as molecular dynamics simulations, we aim to identify the structural and dynamical characteristics of IDPs at the molecular level and correlate them to their biological functions. Building on the knowledge acquired from our previous work, in this grant period, the research will be focused on 1) the mechanisms by which IDPs interact with protein hubs, and 2) the structural and dynamical behaviors of IDPs in crowded environments. Recent studies revealed that a number of well-folded proteins function as critical hubs in the protein-protein interaction network by specifically targeting for IDPs. Elucidating the binding mechanisms between these hubs and IDPs are essential for deciphering the biological functions executed through these protein-protein interactions. Through the extensive studies of the interaction between intrinsically disordered Cby with the 14-3-3 hub protein proposed here, we seek to obtain a better understanding of the factors that govern the affinity and specificity of binding between IDPs and IDP-hubs. Another objective of this proposal is to dissect the effects of macromolecular crowding on IDPs at the molecular level. Structural studies of proteins are usually performed with low concentrations of purified protein in dilute buffer solutions. These conditions, however, are significantly different from that of the cellular environments where proteins carry out their functions. The presence of high concentrations of macromolecules, including protein and DNA, creates a crowded condition in cells. This phenomenon, commonly referred to as molecular crowding, has significant effects on the behaviors of proteins. To investigate the molecular crowding effects on the structure, dynamics, and molecular recognition of IDPs, high concentration of inert agents will be used to mimic the crowded cellular environments and the behaviors of different IDPs with distinct structural characteristics under these conditions will be studied by NMR and other biophysical techniques. The work will shed light on how IDPs functions in crowded cellular environments. Exciting recent experimental discoveries about disordered proteins have generated worldwide research interest in their functions and properties. The results of the proposed research will make a significant contribution to this exciting field.