Impact of Cellular Crowding on Protein Structural Dynamics Investigated by EPR Spectroscopy.

The study of how the intracellular medium influences protein structural dynamics and protein-protein interactions is a captivating area of research for scientists aiming to comprehend biomolecules in their native environment. As the cellular environment can hardly be reproduced , direct investigation of biomolecules within cells has attracted growing interest in the past two decades. Among magnetic resonances, site-directed spin labeling coupled to electron paramagnetic resonance spectroscopy (SDSL-EPR) has emerged as a powerful tool for studying the structural properties of biomolecules directly in cells.

Exploring the dynamics and structure of PpiB in living Escherichia coli cells using electron paramagnetic resonance spectroscopy.

The combined effects of the cellular environment on proteins led to the definition of a fifth level of protein structural organization termed quinary structure. To explore the implication of potential quinary structure for globular proteins, we studied the dynamics and conformations of Escherichia coli (E. coli) peptidyl-prolyl cis/trans isomerase B (PpiB) in E.

investigation of the conformational landscape of the GTPase UreG by SDSL-EPR.

UreG is a cytosolic GTPase involved in the maturation network of urease, an Ni-containing bacterial enzyme. Previous investigations showed that UreG features a flexible tertiary organization, making this protein the first enzyme discovered to be intrinsically disordered. To determine whether this heterogeneous behavior is maintained in the protein natural environment, UreG structural dynamics was investigated directly in intact bacteria by EPR.

Guidelines for the Simulations of Nitroxide X-Band cw EPR Spectra from Site-Directed Spin Labeling Experiments Using S.

Site-directed spin labeling (SDSL) combined with continuous wave electron paramagnetic resonance (cw EPR) spectroscopy is a powerful technique to reveal, at the local level, the dynamics of structural transitions in proteins. Here, we consider SDSL-EPR based on the selective grafting of a nitroxide on the protein under study, followed by X-band cw EPR analysis. To extract valuable quantitative information from SDSL-EPR spectra and thus give a reliable interpretation on biological system dynamics, a numerical simulation of the spectra is required.

Dance with spins: site-directed spin labeling coupled to electron paramagnetic resonance spectroscopy directly inside cells.

Depicting how biomolecules move and interact within their physiological environment is one of the hottest topics of structural biology. This Feature Article gives an overview of the most recent advances in Site-directed Spin Labeling coupled to Electron Paramagnetic Resonance spectroscopy (SDSL-EPR) to study biomolecules in living cells. The high sensitivity, the virtual absence of background, and the versatility of spin-labeling strategies make this approach one of the most promising techniques for the study of biomolecules in physiologically relevant environments.

Probing the Structural Dynamics of a Bacterial Chaperone in Its Native Environment by Nitroxide-Based EPR Spectroscopy.

One of the greatest current challenges in structural biology is to study protein dynamics over a wide range of timescales in complex environments, such as the cell. Among magnetic resonances suitable for this approach, electron paramagnetic resonance spectroscopy coupled to site-directed spin labeling (SDSL-EPR) has emerged as a promising tool to study protein local dynamics and conformational ensembles. In this work, we exploit the sensitivity of nitroxide labels to report protein local dynamics at room temperature.

Nickel and GTP Modulate UreG Structural Flexibility.

UreG is a P-loop GTP hydrolase involved in the maturation of nickel-containing urease, an essential enzyme found in plants, fungi, bacteria, and archaea. This protein couples the hydrolysis of GTP to the delivery of Ni(II) into the active site of apo-urease, interacting with other urease chaperones in a multi-protein complex necessary for enzyme activation. Whereas the conformation of () UreG was solved by crystallography when it is in complex with two other chaperones, in solution the protein was found in a disordered and flexible form, defining it as an intrinsically disordered enzyme and indicating that the well-folded structure found in the crystal state does not fully reflect the behavior of the protein in solution.