Steps of actin filament branch formation by Arp2/3 complex investigated with coarse-grained molecular dynamics.

Abstract

The nucleation of actin filament branches by the Arp2/3 complex involves activation through nucleation promotion factors (NPFs), binding of the complex to the side of actin filaments, and recruitment of actin monomers. Because of the large system size and processes that involve flexible regions and diffuse components, simulations of branch formation using all-atom molecular dynamics are challenging. We applied the Cα based model Kim and Hummer that retains amino-acid level information and allows coarse-grained molecular dynamics (CG-MD) simulations in implicit solvent, with globular domains represented as rigid bodies and flexible regions allowed to fluctuate. We used recent cryo-EM structures of the Arp2/3 complex branch bound to a mother actin filament to represent the activated form of Arp2/3 complex. We studied its interactions with the VCA domain of the NPF Wiskott-Aldrich syndrome protein (WASp) and actin monomers. We found stable configurations with one or two actin monomers bound along the branch filament direction and with VCA domain associated to the strong and weak binding sites of the Arp2/3 complex, supporting prior structural studies and validating our approach. The results further suggest that the interaction between the two actin subunits, and the flexibility of actin D-loop contribute to the stability of the complex. We also performed CG-MD simulations of active Arp2/3 complex bound to a mother actin filament. The bound conformation remained stable during the simulations, which reflect the contribution of each subunit to the binding. D-loop flexibility of actin subunits on the mother filament further stabilized the binding.