Investigating the impact of small molecule ligands and the proteostasis network on protein folding inside the cell

The folded forms of most proteins are critical to their functions. Despite the complexity of the cellular milieu and the presence of high-risk deleterious interactions, there is a high level of fidelity observed in the folding process for entire proteomes. Two important reasons for this are the presence of the quality control machinery consisting of chaperones and degradation enzymes that work jointly to optimize the population of the folded state and interaction partners that re-enforce the functional state and add to the competitive advantage of an organism. While substantial effort has been directed to understand protein folding and interactions in vitro, comparatively little of these processes are explored inside the cell. This work examines two important aspects of protein folding inside the cell; first, the impact of small molecule ligands on protein folding; and second, the impact of the proteostasis network on the folding of an obligatory chaperone client. We deploy a combination of experiments and mathematical modeling based on the principle of kinetic partitioning to understand how these phenomena sculpt the protein folding landscape inside the cell. We find that ligands specifically deplete unfolded and aggregation- or degradation - prone protein populations by favoring the folded state and the chaperone and degradation proteins work to minimize off-pathway species thus reducing the population of aggregated protein inside the cell

Physicochemical Properties of Cells and Their Effects on Intrinsically Disordered Proteins (IDPs)

It has long been axiomatic that a protein’s structure determines its function. Intrinsically disordered proteins (IDPs) and disordered protein regions (IDRs) defy this structure–function paradigm. They do not exhibit stable secondary and/or tertiary structures and exist as dynamic ensembles of interconverting conformers with preferred, nonrandom orientations.(1-4) The concept of IDPs and IDRs as functional biological units was initially met with skepticism. For a long time, disorder, intuitively implying chaos, had no place in our perception of orchestrated molecular events controlling cell biology. Over the past years, however, this notion has changed. Aided by findings that structural disorder constitutes an ubiquitous and abundant biological phenomenon in organisms of all phyla,(5-7) and that it is often synonymous with function,(8-11) disorder has become an integral part of modern protein biochemistry. Disorder thrives in eukaryotic signaling pathways(12) and functions as a prominent player in many regulatory processes.(13-15) Disordered proteins and protein regions determine the underlying causes of many neurodegenerative disorders and constitute the main components of amyloid fibrils.(16) They further contribute to many forms of cancer, diabetes and to cardiovascular and metabolic diseases.(17, 18) Research into disordered proteins produced significant findings and established important new concepts. On the structural side, novel experimental and computational approaches identified and described disordered protein ensembles(3, 19, 20) and led to terms such as secondary structure propensities, residual structural features, and transient long-range contacts.(1, 21) The discovery of coupled folding-and-binding reactions defined the paradigm of disorder-to-order transitions(22) and high-resolution insights into the architectures of amyloid fibrils were obtained.(23, 24) On the biological side, we learned about the unexpected intracellular stability of disordered proteins, their roles in integrating post-translational protein modifications in cell signaling and about their functions in regulatory processes ranging from transcription to cell fate decisions.(15, 25, 26) One open question remaining to be addressed is how these in vitro structural insights relate to biological in vivo effects. How do complex intracellular environments modulate the in vivo properties of disordered proteins and what are the implications for their biological functions (Figure 1)?(27-29