Graduate Program in Biophysics, University of California, San Francisco, San Francisco, CA, USA.
Department of Biochemistry and Biophysics, Institute for Neurodegenerative Diseases, University of California, San Francisco, San Francisco, CA, USA.
Nat Struct Mol Biol. 2020 May;27(5):406-416. doi: 10.1038/s41594-020-0409-5. Epub 2020 Apr 20.
The ClpAP complex is a conserved bacterial protease that unfolds and degrades proteins targeted for destruction. The ClpA double-ring hexamer powers substrate unfolding and translocation into the ClpP proteolytic chamber. Here, we determined high-resolution structures of wild-type Escherichia coli ClpAP undergoing active substrate unfolding and proteolysis. A spiral of pore loop-substrate contacts spans both ClpA AAA+ domains. Protomers at the spiral seam undergo nucleotide-specific rearrangements, supporting substrate translocation. IGL loops extend flexibly to bind the planar, heptameric ClpP surface with the empty, symmetry-mismatched IGL pocket maintained at the seam. Three different structures identify a binding-pocket switch by the IGL loop of the lowest positioned protomer, involving release and re-engagement with the clockwise pocket. This switch is coupled to a ClpA rotation and a network of conformational changes across the seam, suggesting that ClpA can rotate around the ClpP apical surface during processive steps of translocation and proteolysis.
ClpAP 复合物是一种保守的细菌蛋白酶，可展开并降解靶向破坏的蛋白质。ClpA 双环六聚体为底物展开和易位到 ClpP 蛋白水解腔提供动力。在这里，我们确定了处于主动底物展开和蛋白水解状态的野生型大肠杆菌 ClpAP 的高分辨率结构。孔环-底物接触的螺旋横跨两个 ClpA AAA+ 结构域。螺旋缝处的原体经历核苷酸特异性重排，支持底物易位。IGL 环灵活延伸，与平面的七聚体 ClpP 表面结合，空的、对称性不匹配的 IGL 口袋在缝处保持。通过最低位置原体的 IGL 环识别三种不同的结构，涉及与顺时针口袋的释放和重新结合。这种开关与 ClpA 的旋转以及缝处的构象变化网络耦合在一起，表明 ClpA 可以在易位和蛋白水解的连续步骤中绕 ClpP 顶端表面旋转。
