STRUCTURE AND FUNCTION OF THE CLPXP ATP-DEPENDENT PROTEASE

CLPXP ATP 依赖性蛋白酶的结构和功能

• 批准号: 7369492
• 负责人: TANIA A BAKER
• 金额: $ 0.07万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-06-01 至 2007-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7369492
• 关键词: STRUCTURE; FUNCTION; CLPXP; ATP; DEPENDENT

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. We had previously solved the structure of the AAA+ adaptor protein SspB in complex with one of its substrates, the ssrA tag. In E. coli, this tag targets incompletely synthesized proteins from stalled ribosomes for destruction by the ClpXP degradation machine, a process that is accelerated by SspB. Biochemical studies have revealed that other substrates are targeted to ClpXP by SspB, including the n-terminal fragment of RseA. RseA is a trans-membrane protein that functions as a master regulator of the extra-cytoplasmic stress response in E. coli. Targeting of n-terminal fragment for degradation by ClpXP is required for the release of the sigma factor (sigma E) and activation stress response genes. Using biochemical and biophysical techniques, a 31 amino acid stretch of the RseA sequence was identified as the SspB-binding determinant. In order to understand the structural basis for the SspB-RseA interaction, we crystallized the protein in complex with this 31 amino acid peptide. Energy-dependent proteases often rely on adaptor proteins to modulate substrate recognition. The SspB adaptor binds peptide sequences in the stress-response regulator RseA and in ssrA-tagged proteins and delivers these molecules to the AAA+ ClpXP protease for degradation. The structure of SspB bound to an ssrA peptide is known. Here, we reported the crystal structure of a complex between SspB and its recognition peptide in RseA in Nat. Struct. Mol. Biol. Notably, the RseA sequence is positioned in the peptide-binding groove of SspB in a direction opposite to the ssrA peptide, the two peptides share only one common interaction with the adaptor, and the RseA interaction site is substantially larger than the overlapping ssrA site. This marked diversity in SspB recognition of different target proteins indicates that it is capable of highly flexible and dynamic substrate delivery

该子项目是利用 NIH/NCRR 资助的中心拨款提供的资源的众多研究子项目之一。子项目和研究者 (PI) 可能已从另一个 NIH 来源获得主要资金，因此可以在其他 CRISP 条目中出现。列出的机构是中心的机构，不一定是研究者的机构。我们之前已经解决了 AAA+ 接头蛋白 SspB 与其底物之一 ssrA 标签复合的结构。在大肠杆菌中，该标签以停滞核糖体中不完全合成的蛋白质为目标，并通过 ClpXP 降解机进行破坏，这一过程由 SspB 加速。生化研究表明，SspB 还可以靶向 ClpXP 的其他底物，包括 RseA 的 n 末端片段。 RseA 是一种跨膜蛋白，在大肠杆菌中充当胞质外应激反应的主要调节因子。释放 sigma 因子 (sigma E) 和激活应激反应基因需要 ClpXP 靶向 N 末端片段进行降解。使用生物化学和生物物理技术，RseA 序列的 31 个氨基酸序列被鉴定为 SspB 结合决定簇。为了了解 SspB-RseA 相互作用的结构基础，我们结晶了与该 31 个氨基酸肽复合的蛋白质。能量依赖性蛋白酶通常依靠衔接蛋白来调节底物识别。 SspB 接头结合应激反应调节剂 RseA 和 ssrA 标记蛋白中的肽序列，并将这些分子传递至 AAA+ ClpXP 蛋白酶进行降解。与 ssrA 肽结合的 SspB 的结构是已知的。在这里，我们报道了 Nat 中 RseA 中 SspB 与其识别肽之间的复合物的晶体结构。结构。摩尔。生物。值得注意的是，RseA序列位于SspB的肽结合沟中，方向与ssrA肽相反，这两个肽仅与接头有一种共同的相互作用，并且RseA相互作用位点比重叠的ssrA位点大得多。 SspB 识别不同靶蛋白的显着多样性表明它能够高度灵活和动态的底物递送