Concerted release of substrate domains from GroEL by ATP is demonstrated with FRET.
Concerted release of substrate domains from GroEL by ATP is demonstrated with FRET.
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FRET 证明了 ATP 从 GroEL 中协调释放底物结构域。
DOI：
10.1016/j.jmb.2008.05.021
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发表时间：
2008
影响因子：
5.6
通讯作者：
Horovitz,Amnon
中科院分区：
文献类型：
--
作者：
Papo,Niv;Kipnis,Yakov;Haran,Gilad;Horovitz,Amnon
伴侣蛋白GroEL通过在其两个背靠背堆叠的环中的每一个内进行ATP诱导的构象变化来协助蛋白质折叠。在这里，我们研究了是否协调变构开关引起全或无释放和折叠的结构域在嵌合荧光蛋白底物，CyPet-YPet。使用这种底物，可以通过测量两个结构域之间的Förster共振能量转移，从其固有荧光和整个嵌合体确定每个结构域的折叠产率。因此，可以确定一个域的释放是否伴随着另一个域的释放（协调机制），或者它们的释放是否不耦合。我们的研究结果表明，嵌合体的释放往往是一致的折叠时，由野生型GroEL变体的辅助，但不是当辅助的F44 W/D155 A突变体，经历了顺序变构开关。这种分子机器的变构机制与其辅助折叠的生物学功能之间的联系由此建立。
The chaperonin GroEL assists protein folding by undergoing ATP-induced conformational changes that are concerted within each of its two back-to-back stacked rings. Here we examined whether concerted allosteric switching gives rise to all-or-none release and folding of domains in a chimeric fluorescent protein substrate, CyPet–YPet. Using this substrate, it was possible to determine the folding yield of each domain from its intrinsic fluorescence and that of the entire chimera by measuring Förster resonance energy transfer between the two domains. Hence, it was possible to determine whether release of one domain is accompanied by release of the other domain (concerted mechanism), or whether their release is not coupled. Our results show that the chimera's release tends to be concerted when folding is assisted by a wild-type GroEL variant, but not when assisted by the F44W/D155A mutant that undergoes a sequential allosteric switch. A connection between the allosteric mechanism of this molecular machine and its biological function in assisting folding is thus established.