Structure and Activity of ClpB from Escherichia coli
Structure and Activity of ClpB from Escherichia coli
复制标题
大肠杆菌 ClpB 的结构和活性
DOI：
--
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发表时间：
2000
影响因子：
4.8
通讯作者：
M. Żółkiewski
中科院分区：
文献类型：
--
作者：
Micheal E. Barnett;A. Zolkiewska;M. Żółkiewski
ClpB是大肠杆菌中蛋白解聚多分子伴侣系统的成员。ClpB介导的蛋白质折叠反应的机制目前尚不清楚，ClpB中不同序列区域的功能作用正在讨论中。我们已经表达并纯化了全长ClpB和三个截短的变体，其具有N-末端、C-末端和双N-和C-末端缺失。我们研究了蛋白浓度依赖性和ATP诱导的ClpB寡聚化，酪蛋白诱导的ClpB ATP酶激活，和ClpB辅助变性萤火虫荧光素酶的再激活。我们发现ClpB的N-和C-末端截短强烈抑制其分子伴侣活性。这种抑制的原因是不同的，但是，对于N-和C-末端截短。C-末端结构域的缺失抑制ClpB的自缔合，这导致对ATP的亲和力降低，并且导致C-末端截短的变体的ATP酶和伴侣活性降低。与此相反，N-末端结构域的缺失并不抑制ClpB的自缔合及其基础ATP酶活性，但降低了酪蛋白激活ClpB ATP酶的能力。这些结果表明，N-末端区域的ClpB可能含有功能上重要的蛋白结合位点，而C-末端区域的主要作用是支持寡聚化的ClpB。
ClpB is a member of a protein-disaggregating multi-chaperone system in Escherichia coli. The mechanism of protein-folding reactions mediated by ClpB is currently unknown, and the functional role of different sequence regions in ClpB is under discussion. We have expressed and purified the full-length ClpB and three truncated variants with the N-terminal, C-terminal, and a double N- and C-terminal deletion. We studied the protein concentration-dependent and ATP-induced oligomerization of ClpB, casein-induced activation of ClpB ATPase, and ClpB-assisted reactivation of denatured firefly luciferase. We found that both the N- and C-terminal truncation of ClpB strongly inhibited its chaperone activity. The reasons for such inhibition were different, however, for the N- and C-terminal truncation. Deletion of the C-terminal domain inhibited the self-association of ClpB, which led to decreased affinity for ATP and to decreased ATPase and chaperone activity of the C-terminally truncated variants. In contrast, deletion of the N-terminal domain did not inhibit the self-association of ClpB and its basal ATPase activity but decreased the ability of casein to activate ClpB ATPase. These results indicate that the N-terminal region of ClpB may contain a functionally significant protein-binding site, whereas the main role of the C-terminal region is to support oligomerization of ClpB.
DOI：
10.1016/s0021-9258(17)41804-7
发表时间：
1994-02
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
D. Parsell;A. Kowal;S. Lindquist
通讯作者：
D. Parsell;A. Kowal;S. Lindquist
影响因子：
2.9
作者：
STAFFORD, WF
通讯作者：
STAFFORD, WF