Dimerization of the human E3 ligase CHIP via a coiled-coil domain is essential for its activity
Dimerization of the human E3 ligase CHIP via a coiled-coil domain is essential for its activity
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DOI：
10.1074/jbc.m311112200
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发表时间：
2004-01-23
影响因子：
4.8
通讯作者：
Bukau, B
中科院分区：
文献类型：
--
作者：
Nikolay, R;Wiederkehr, T;Bukau, B
热休克蛋白70相互作用的E3-泛素连接酶CHIP已被牵连在决定是否进入重折叠或降解途径的靶蛋白。为了进一步表征CHIP的活性，我们纯化了未标记的智人和黑腹果蝇CHIP（hCHIP，dCHIP）。与其他E3-泛素连接酶相比，hCHIP和dCHIP蛋白在生理浓度下形成同源二聚体。我们确定了一个预测的卷曲螺旋区域中的混合电荷段的hCHIP和dCHIP序列，并发现它是必要的和足够的二聚体形成。缺少该片段的hCHIP突变体（hCHIP Δ（128-229））不能形成二聚体，但该片段本身（hCHIP-（128-229））容易二聚化。此外，我们证明了二聚化是hCHIP在重构泛素化测定中的活性的先决条件。因此，二聚化的控制可以提供用于调节CHIP活性的机制。
The Hsp70-interacting E3-ubiquitin ligase CHIP has been implicated in the decision as to whether a target protein enters the refolding or the degradation pathway. To further characterize the activity of CHIP we purified untagged Homo sapiens and Drosophila melanogaster CHIP (hCHIP, dCHIP). In contrast to other E3-ubiquitin ligases, both hCHIP and dCHIP proteins formed homodimers at physiological concentrations. We identified a predicted coiled-coil region in a mixed charge segment of the hCHIP and dCHIP sequence and found it to be necessary and sufficient for dimer formation. A mutant of hCHIP lacking this segment (hCHIPDelta(128-229)) was incapable of dimer formation, but the segment by itself (hCHIP-(128-229)) readily dimerized. Furthermore, we demonstrated that dimerization is a prerequisite for activity of hCHIP in the reconstituted ubiquitination assay. Control of dimerization may thus provide a mechanism for regulation of CHIP activity.