Actin is an abundant and essential cytoskeletal protein that plays a central role in eukaryotic cell structure and functions. The assembly of actin monomers into dynamic filaments is important for many cellular processes including cell division, cell movement, force generation, and muscle contraction. The intracellular environment where actin assembly occurs is crowded with various organelles, proteins, and macromolecules that limit the accessible volume for biomolecular interactions and reactions. Previous studies have shown the effects of macromolecular crowding on actin filament assembly. However, the impact of crowder size on actin assembly dynamics and kinetics is not well understood.  In this study we investigate how the excluded volume effect caused by crowding influences actin filament assembly dynamics by using synthetic polymeric crowder, polyethylene glycol (PEG), of various molecular weights. Direct visualization of filament assembly in the presence of PEG is achieved by total internal reflection fluorescence (TIRF) microscopy imaging. Furthermore, the effects of crowder size on actin polymerization kinetics were evaluated by pyrene fluorescence assays. We demonstrate that the molecular weight of PEG modulates the steady-state average filament lengths as well as the filament assembly rates. Taken together, our data suggests that the size of macromolecular crowders impacts actin assembly dynamics and kinetics<i> in vitro</i>.

Effects of crowder size on actin assembly dynamics and kinetics

ACS Spring Meeting 2021

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