Control of Substrate Gating and Translocation into ClpP by Channel Residues and ClpX Binding
Control of Substrate Gating and Translocation into ClpP by Channel Residues and ClpX Binding
复制标题
DOI：
10.1016/j.jmb.2010.04.027
复制
发表时间：
2010-06-25
影响因子：
5.6
通讯作者：
Sauer, Robert T.
中科院分区：
文献类型：
--
作者：
Lee, Mary E.;Baker, Tania A.;Sauer, Robert T.
ClpP是自区室化蛋白酶，其具有非常有限的降解活性，除非其与ClpX缔合以形成ClpXP或与ClpA缔合以形成ClpAP。在这里，我们表明，ClpX结合刺激ClpP裂解的肽大于几个氨基酸，并增强ClpP活性位点的修饰。刺激需要ATP结合，但不需要ClpX水解。这种增强的幅度与进入ClpP的分子的分子量增加相关。ClpP的通道环或螺旋A中的氨基酸取代增强较大底物进入游离酶，在某些情况下消除ClpX结合，并且在其他情况下不被ClpX结合进一步刺激。这些结果支持一种模型，其中游离ClpP的通道残基排除了除最小肽外的所有肽有效进入降解室，ClpX结合用于缓解这些抑制性相互作用。特异性ClpP通道变体也阻止某些氨基酸序列的ClpXP易位，表明野生型通道在促进广泛的易位特异性中起重要作用。结合以前的研究，我们的研究结果表明，ClpP和它的合作伙伴ATP酶之间的合作打开了一扇大门，其功能是从孤立的ClpP排除较大的基板。(C)2010爱思唯尔有限公司版权所有。
ClpP is a self-compartmentalized protease, which has very limited degradation activity unless it associates with ClpX to form ClpXP or with ClpA to form ClpAP. Here, we show that ClpX binding stimulates ClpP cleavage of peptides larger than a few amino acids and enhances ClpP active-site modification. Stimulation requires ATP binding but not hydrolysis by ClpX. The magnitude of this enhancement correlates with increasing molecular weight of the molecule entering ClpP. Amino-acid substitutions in the channel loop or helix A of ClpP enhance entry of larger substrates into the free enzyme, eliminate ClpX binding in some cases, and are not further stimulated by ClpX binding in other instances. These results support a model in which the channel residues of free ClpP exclude efficient entry of all but the smallest peptides into the degradation chamber, with ClpX binding serving to relieve these inhibitory interactions. Specific ClpP channel variants also prevent ClpXP translocation of certain amino-acid sequences, suggesting that the wild-type channel plays an important role in facilitating broad translocation specificity. In combination with previous studies, our results indicate that collaboration between ClpP and its partner ATPases opens a gate that functions to exclude larger substrates from isolated ClpP. (C) 2010 Elsevier Ltd. All rights reserved.