Welcome to a LINXS-ECIS webinar with Daniel Harries (The Hebrew University of Jerusalem) and Gary Pielak (University of North Carolina)
When: 4 February, 15:00 - 17:00
Where: Zoom - participants will receive a link the same day upon registration
Title: Crowding beyond excluded volume: a tale of two dimers
Speakers: Daniel Harries, Professor of Chemistry, The Hebrew University of Jerusalem; and Gary Pielak, Kenan Distinguished Professor of Chemistry, Biochemistry and Biophysics , University of North Carolina
Abstract
Protein-protein interactions are modulated by their environment. High macromolecular solute concentrations crowd proteins and shift equilibria between protein monomers and their assemblies. We aim to understand the mechanism of crowding by elucidating the molecular-level interactions that determine dimer stability. Using 19F-NMR spectroscopy, we studied the effects of various polyethylene glycols (PEGs) on the equilibrium thermodynamics of two protein complexes: a side-by-side and a domain-swap dimer. Analysis using our mean-field crowding model shows that, contrary to classic crowding theories, PEGs destabilize both dimers through enthalpic interactions between PEG and the monomers. The enthalpic destabilization becomes more dominant with increasing PEG concentration, because the reduction in PEG mesh size with concentration diminishes the stabilizing effect of excluded volume interactions. Additionally, the partially folded domain-swap monomers fold in the presence of PEG, contributing to dimer destabilization at low PEG concentrations. Our results reveal that polymers crowd protein complexes through multiple conjoined mechanisms, impacting both their stability and oligomeric state.
Biographies
Daniel Harries has made significant advances in understanding how biologically diverse environments influence macromolecular behavior through research at the interface of physical chemistry and molecular biophysics. Harries has developed theoretical frameworks, closely integrated with experimental findings, to dissect the forces driving macromolecular associations, dissociations, and functional complex formation in cells. His work focuses on systems such as protein folding, association, and aggregation, and lipid membrane properties and interactions.
A key area of Harries’s research explores the effects of molecular crowding, osmotic pressures, and confinement on macromolecular stability. By addressing processes such as peptide folding and aggregation into amyloid fibers, membrane confinement in nanodiscs, and viral assembly, Harries has provided new insights into how macromolecular systems function in realistic cellular environments. His work underscores how forces such as depletion interactions, crowding, and confinement play important roles in shaping biological processes, with broad implications for health and disease.
Gary Pielak has made fundamental discoveries in the field of protein chemistry through highly original research to unravel protein biophysics undercrowded conditions in vitro and in living cells. Pielak has developed innovative and quantitative techniques for measuring protein structure, stability, diffusion, and concentration in living cells and under crowded in vitro conditions. His efforts have led to major advances in understanding how the intracellular environment impacts both globular and intrinsically disordered proteins. Prior to his pioneering efforts, almost all knowledge about proteins came from studies under artificial environments involving either dilute buffer or solutions crowded with synthetic polymers. Pielak’s work has revolutionized our understanding of how proteins work where they actually function – inside cells. Using in-cell nuclear magnetic resonance spectroscopy, a technique he helped develop, his pioneering work has overturned the decades-old idea that the impacts of crowding arise solely from the close packed nature of the cytoplasm. Instead, his work shows that repulsive and attractive chemical interactions between cellular components determine the effect of macromolecular crowding and has recently presented a quantitative model to explain crowding effects that is independent of crowder identity. These interactions organize the inside of cells, controlling metabolism and signaling. His research reveals the true consequences of the cellular environment on proteins and creates new opportunities for physiologically relevant biophysics.
ContacT
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