Conformational Changes Of Arp2/3 Complex During Activation

Abstract

Kinetic and structural studies of Actin-related protein (Arp) 2/3 complex point to a transition from inactive to active conformations at an actin branch point. The inactive conformation is greatly favored for purified Arp2/3 complex. Crystal structures of inactive Arp2/3 complex show that Arp2 and Arp3 are separated by about 3 nm compared to their positions as the first two subunits of the daughter filament in low resolution models based in reconstructions from electron tomograms of actin filament branch junctions. Here, we use atomistic-scale molecular dynamics simulations to study activation of Arp2/3 complex. Starting in the ATP-bound inactive crystal structure, we apply forces to Arp2 and Arp3 so that the energy of the system is at a minimum when Arp2 and Arp3 are positioned like the first and second subunits of the daughter filament. Arp2 does not detach from the other subunits during activation as proposed from biochemical experiments. The long C-terminal α-helix of p34 rotates along with Arp2 and p40 through interactions with p20. Residues His30-Leu35 of p20 stay in contact with the residues of the αG helix of Arp2 throughout the course of the simulation. Also, residues Arg106-Lys107 of p20 interact with the region of Arp2 surrounding Leu235 between the αF and αG loops. The main impediment to a smooth transition is the αK helix of subdomain 3 of Arp3, which collides with the DNase binding loop of Arp2. This study provides the first insights into the conformational changes of Arp2/3 complex forms an actin filament branch.