Project: 4 Molecular dynamics simulations to analyze the dynamics of Arp2/3 cmplx

肌动蛋白丝分支连接的3D结构，从电子分辨率为2.5 nm 断层扫描证实Arp 2位于分支连接处Arp 3旁边，与Arp形成一致 短节距肌动蛋白丝螺旋（5）。我们的PPG合作者将非活性晶体结构放入 通过Arp 2的刚体运动紧接Arp 3的断层摄影密度。由于可以分离Arp 2/3复合物， 缺少Arp 2（S2），可以想象Arp 2以这种方式移动，通过N末端与复合物的其余部分相连 ARPC5。然而，我们也发现，裂殖酵母存活ARPC 5的N-末端的删除 与正常出现的肌动蛋白斑块（诺伦和波拉德，未发表的意见）。因此我们要 考虑其他机制来将Arp 2移动到Arp 3旁边，例如保持Arp 2与ARPC 1的接触， ARPC 2和ARPC 4，同时弯曲ARPC 2/和ARPC 4（以及其他子单元）以使Arp 2靠近Arp 3。 基本原理：我们认为整个分支结的结晶不太可能，但分子动力学 模拟提供了一种替代的实验方法来探索构象变化，发生在 Arp 2/3复合物在从非活动状态到分支连接处的转变过程中。

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that fission yeast survive deletion of the N-terminus of ARPC5 with normal appearing actin patches (Nolen and Pollard, unpublished observations).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.