Binding of the ClpA Unfoldase Opens the Axial Gate of ClpP Peptidase
Binding of the ClpA Unfoldase Opens the Axial Gate of ClpP Peptidase
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DOI：
10.1074/jbc.m109.090498
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发表时间：
2010-05-07
影响因子：
4.8
通讯作者：
Steven, Alasdair C.
中科院分区：
文献类型：
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作者：
Effantin, Gregory;Maurizi, Michael R.;Steven, Alasdair C.
ClpP是一种丝氨酸蛋白酶，其活性位点被隔离在两个七聚体亚基环之间的空腔中。ClpP处理折叠蛋白质底物的能力取决于其与AAA+ ATP酶/解折叠酶、ClpA或ClpX配对。在活性复合物中，底物被展开并沿着轴向通道进料至ClpP内的降解室。我们已经使用冷冻电子显微镜在类似于11埃的分辨率，以调查的三维结构的ClpP复杂的一个或两个端部安装ClpA六聚体。在不存在ClpA的情况下，ClpP的顶端区域是密封的;然而，它在ClpA结合时打开，产生进入通道。该区域被ClpP的N-末端环（残基1-17）占据，其在晶体结构中往往难以看到，指示构象可变性。尽管如此，我们能够模拟伴随ClpA结合这些环的运动的封闭到开放的转变;特别是，“向上”的环构象与开放状态相关。ClpP的主要部分，即由残基18-193的14个拷贝形成的桶，基本上不受与ClpA的相互作用的影响。使用差异映射，我们本地化的结合位点ClpA相邻ClpP亚基之间的外周口袋。基于这些观察结果，我们提出，进入ClpP降解室是由其N-末端环的铰链运动变构控制的，其中ClpA的不匹配结合足以诱导。
ClpP is a serine protease whose active sites are sequestered in a cavity enclosed between two heptameric rings of subunits. The ability of ClpP to process folded protein substrates depends on its being partnered by an AAA+ ATPase/unfoldase, ClpA or ClpX. In active complexes, substrates are unfolded and fed along an axial channel to the degradation chamber inside ClpP. We have used cryoelectron microscopy at similar to 11-angstrom resolution to investigate the three-dimensional structure of ClpP complexed with either one or two end-mounted ClpA hexamers. In the absence of ClpA, the apical region of ClpP is sealed; however, it opens on ClpA binding, creating an access channel. This region is occupied by the N-terminal loops (residues 1-17) of ClpP, which tend to be poorly visible in crystal structures, indicative of conformational variability. Nevertheless, we were able to model the closed-to-open transition that accompanies ClpA binding in terms of movements of these loops; in particular, "up" conformations of the loops correlate with the open state. The main part of ClpP, the barrel formed by 14 copies of residues 18-193, is essentially unchanged by the interaction with ClpA. Using difference mapping, we localized the binding site for ClpA to a peripheral pocket between adjacent ClpP subunits. Based on these observations, we propose that access to the ClpP degradation chamber is controlled allosterically by hinged movements of its N-terminal loops, which the symmetry-mismatched binding of ClpA suffices to induce.