Investigating the effects of molecular crowding on the kinetics of protein aggregation

Abstract

The thermodynamics and kinetics of protein folding and protein aggregation in vivo are of great importance in numerous scientific areas including fundamental biophysics research, nanotechnology, and medicine. However, these processes remain poorly understood in both in vivo and in vitro systems. Here we extend an established model for protein aggregation that is based on the kinetic equations for the moments of the polymer size distribution by introducing macromolecular crowding particles into the model using scaled-particle and transition-state theories. The model predicts that the presence of crowders can either speed up, cause no change to, or slow down the progress of the aggregation compared to crowder-free solutions, in striking agreement with experimental results from nine different amyloid-forming proteins that utilized dextran as the crowder. These different dynamic effects of macromolecular crowding can be understood in terms of the change of excluded volume associated with each reaction step.

Response to crowded conditions reveals compact nucleus for amyloid formation of folded protein

“…This is in excellent agreement with excluded volume theory that predicts that assembled species are favored when the volume is reduced (Minton, 2005b;Ellis and Minton, 2006). Also more elaborate modelling (including both scaled-particle and transition-state theories) predicts this for systems where the size of the monomer is smaller when in the amyloid fiber than when in the free form (Schreck et al, 2020). Notably, 200 mg ml À1 Ficoll 70 will occupy 13% of the volume based on its partial specific volume value (Christiansen et al, 2010;Aguilar et al, 2011) and thus the effective concentration of apo-β-PV is not 30 μM but 34 μM in the presence of 200 mg ml À1 Ficoll 70.…”

“…In addition to excluded volume and solvation effects (studied here), also nonspecific chemical interactions may take place between macromolecules and hydrophobic forces within macromolecules, for example, DNA, were shown to be affected by cell-like conditions (Feng et al, 2019). Many more systematic studies of these effects on the molecularmechanistic level are desired, including studies using mixtures of macromolecular crowding agents, protein crowders, as well as assessing combinations of macromolecular crowding agents and osmolytes; all in combination with new theoretical approaches (Schreck et al, 2020). So far, known amyloidogenic proteins involved in human diseases are intrinsically disordered proteins or folded (often oligomeric) proteins that require partial unfolding/ dissociation (i.e.…”

Virtual Issue on Protein Crowding and Stability

“…27 This was interpreted to result from SOD1− crowder interactions that were competing with SOD1 aggregation as one example of crowding-induced effects on aggregation covered more generally in a theoretical analysis. 28 Homing in on biological function, a FRET-based analysis contrasts binding of Hsp70 and Hsc70 to a model protein substrate in living cells in the cellular background of other functional interactions of Hsc70. 29 Interesting related work, although not strictly under crowded conditions, illustrates for KRAS how its functional interactions are modulated by ubiquitination.…”

Probing the role of macromolecular crowding in cell volume regulation using fluorescence anisotropy

“…Consequently, the fraction of excluded volume of the macromolecules becomes notable compared to the total volume of the cytoplasm. This entropic effect is also described as the thermodynamic state of macromolecular crowding (MMC) (Minton, 1981; Ellis, 2001), which affects a host of physicochemical processes in the cytoplasm such as diffusion (Rashid et al ., 2015), active transport (Nettesheim et al ., 2020), protein-ligand binding kinetics (Minton, 2001; Köhn et al ., 2021), enzyme-substrate reaction rates (Thoke, Bagatolli and Olsen, 2018; Wilcox, Chung and Slade, 2021), macromolecular self-assembly (André and Spruijt, 2020; Schreck, Bridstrup and Yuan, 2020), protein folding (Ådén and Wittung-Stafshede, 2014) and post-translational modifications (Darling and Uversky, 2018), to name a few. Consequently, it led to a hypothesis that cells may regulate their cytoplasm MMC to control biochemical processes.…”