School of Life Sciences and Biotechnology, Korea University, Seoul 136-701, Korea.
Mol Cells. 2011 Dec;32(6):589-95. doi: 10.1007/s10059-011-0197-1. Epub 2011 Nov 9.
ClpP is a cylindrical protease that is tightly regulated by Clp-ATPases. The activation mechanism of ClpP using acyldepsipeptide antibiotics as mimics of natural activators showed enlargement of the axial entrance pore for easier processing of incoming substrates. However, the elimination of degradation products from inside the ClpP chamber remains unclear since there is no exit pore for releasing these products in all determined ClpP structures. Here we report a new crystal structure of ClpP from Bacillus subtilis, which shows a significantly compressed shape along the axial direction. A portion of the handle regions comprising the heptameric ring-ring contacts shows structural transition from an ordered to a disordered state, which triggers the large conformational change from an extended to an overall compressed structure. Along with this structural change, 14 side pores are generated for product release and the catalytic triad adopts an inactive orientation. We have also determined B. subtilis ClpP inhibited by diisopropylfluoro-phosphate and analyzed the active site in detail. Structural information pertaining to several different conformational steps such as those related to extended, ADEP-activated, DFP-inhibited and compressed forms of ClpP from B. subtilis is available. Structural comparisons suggest that functionally important regions in the ClpP-family such as N-terminal segments for the axial pore, catalytic triads, and handle domains for the product releasing pore exhibit intrinsically dynamic and unique structural features. This study provides valuable insights for understanding the enigmatic cylindrical degradation machinery of ClpP as well as other related proteases such as HslV and the 20S proteasome.
ClpP 是一种圆柱形蛋白酶，其活性受到 Clp-ATPases 的严格调控。利用 acyldepsipeptide 抗生素作为天然激活剂的模拟物来研究 ClpP 的激活机制表明，轴向入口孔的扩大使得进入的底物更容易被加工。然而，由于在所有已确定的 ClpP 结构中都没有出口孔来释放这些产物，因此仍然不清楚如何从 ClpP 腔室内部清除降解产物。在这里，我们报告了枯草芽孢杆菌 ClpP 的一个新晶体结构，该结构显示沿轴向方向显著压缩的形状。由七聚体环-环接触组成的手柄区域的一部分显示出从有序到无序状态的结构转变，这引发了从扩展到整体压缩结构的大构象变化。伴随着这种结构变化，产生了 14 个侧孔以释放产物，并且催化三联体采用非活性取向。我们还确定了被二异丙基氟磷酸抑制的枯草芽孢杆菌 ClpP，并详细分析了活性位点。与来自枯草芽孢杆菌的伸展、ADEP 激活、DFP 抑制和压缩形式相关的几个不同构象步骤的结构信息都可获得。结构比较表明，ClpP 家族中功能重要的区域，如轴向孔的 N 端片段、催化三联体和用于产物释放孔的手柄结构域，具有固有动态和独特的结构特征。这项研究为理解 ClpP 这一神秘的圆柱形降解机制以及其他相关蛋白酶，如 HslV 和 20S 蛋白酶体，提供了有价值的见解。
