The 3D structure of the actin filament branch junction at 2.5 nm resolution from electron tomography confirmed that Arp2 is located next to Arp3 in the branch junction, consistent with the Arps forming a short-pitch actin filament helix (5). Our PPG collaborators fit the inactive crystal structure into the tomographic density by a rigid body movement of Arp2 next to Arp3. Since one can isolate Arp2/3 complex lacking Arp2 (S2), it is conceivable that Arp2 moves in this way, tethered to the rest of the complex by the Nterminus of ARPC5. However, we also found that fission yeast survive deletion of the N-terminus of ARPC5 with normal appearing actin patches (Nolen and Pollard, unpublished observations). Therefore, we must consider other mechanisms to move Arp2 next to Arp3, such as maintaining the contacts of Arp2 with ARPC1, ARPC2 and ARPC4 while bending ARPC2/and ARPC4 (and other subunits) to bring Arp2 next to Arp3. Rationale: We feel that crystallization of the entire branch junction is unlikely, but molecular dynamics simulations offer an alternative experimental approach to explore the conformation change that occurs within Arp2/3 complex during the transition from the inactive state to the branch junction.

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Volkmann, Niels (2014) The joys and perils of flexible fitting. Adv Exp Med Biol 805:137-55
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