Single molecule techniques are particularly well suited for investigating the processes of protein folding and chaperone assistance. However, current assays provide only a limited perspective on... Show moreSingle molecule techniques are particularly well suited for investigating the processes of protein folding and chaperone assistance. However, current assays provide only a limited perspective on the various ways in which the cellular environment can influence the folding pathway of a protein. In this study, a single molecule mechanical interrogation assay is developed and used to monitor protein unfolding and refolding within a cytosolic solution. This allows to test the cumulative topological effect of the cytoplasmic interactome on the folding process. The results reveal a stabilization against forced unfolding for partial folds, which are attributed to the protective effect of the cytoplasmic environment against unfolding and aggregation. This research opens the possibility of conducting single molecule molecular folding experiments in quasi-biological environments. Show less
Biological polymers, including proteins and the genome, undergo folding processes crucial for their proper functioning. Even slight changes in the folding structure of these biopolymers can have... Show moreBiological polymers, including proteins and the genome, undergo folding processes crucial for their proper functioning. Even slight changes in the folding structure of these biopolymers can have significant implications, leading to the development of various pathological conditions, such as neurodegenerative diseases and cancer. In this thesis, we leverage the theoretical framework of Circuit Topology and expand its application to real-world scenarios. By employing this approach, we quantify the folding patterns of biological polymers, offering valuable insights for detecting harmful misfolds. Furthermore, this research holds the potential to provide fundamental design principles for molecular engineering in the realm of pharmaceutical applications. Show less
Scalvini, B.; Sheikhhassani, V.; Brug, N. van de; Heling, L.W.H.J.; Schmit, J.D.; Mashaghi Tabari, A. 2023
Intrinsically disordered proteins (IDPs) lack a stable native conformation, making it challenging to characterize their structure and dynamics. Key topological motifs with fundamental biological... Show moreIntrinsically disordered proteins (IDPs) lack a stable native conformation, making it challenging to characterize their structure and dynamics. Key topological motifs with fundamental biological relevance are often hidden in the conformational noise, eluding detection. Here, we develop a circuit topology toolbox to extract conformational patterns, critical contacts, and timescales from simulated dynamics of intrinsically disordered proteins. We follow the dynamics of IDPs by providing a smart low-dimensionality representation of their three-dimensional (3D) configuration in the topology space. Such an approach allows us to quantify topological similarity in dynamic systems, therefore providing a pipeline for structural comparison of IDPs. Show less
What is the topology of a protein and what governs protein folding to a specific topology? This is a fundamental question in biology. The protein folding reaction is a critically important cellular... Show moreWhat is the topology of a protein and what governs protein folding to a specific topology? This is a fundamental question in biology. The protein folding reaction is a critically important cellular process, which is failing in many prevalent diseases. Understanding protein folding is also key to the design of new proteins for applications. However, our ability to predict the folding of a protein chain is quite limited and much is still unknown about the topological principles of folding. Current predictors of folding kinetics, including the contact order and size, present a limited predictive power, suggesting that these models are fundamentally incomplete. Here, we use a newly developed mathematical framework to define and extract the topology of a native protein conformation beyond knot theory, and investigate the relationship between native topology and folding kinetics in experimentally characterized proteins. We show that not only the folding rate, but also the mechanistic insight into folding mechanisms can be inferred from topological parameters. We identify basic topological features that speed up or slow down the folding process. The approach enabled the decomposition of protein 3D conformation into topologically independent elementary folding units, called circuits. The number of circuits correlates significantly with the folding rate, offering not only an efficient kinetic predictor, but also a tool for a deeper understanding of theoretical folding models. This study contributes to recent work that reveals the critical relevance of topology to protein folding with a new, contact-based, mathematically rigorous perspective. We show that topology can predict folding kinetics when geometry-based predictors like contact order and size fail. Show less
