Fast-scanning atomic force microscopy reveals the ATP/ADP-dependent conformational changes of GroEL
Fast-scanning atomic force microscopy reveals the ATP/ADP-dependent conformational changes of GroEL
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DOI：
10.1038/sj.emboj.7601326
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发表时间：
2006-10-04
期刊：
影响因子：
11.4
通讯作者：
Takeyasu, Kunio
中科院分区：
文献类型：
--
作者：
Yokokawa, Masatoshi;Wada, Chieko;Takeyasu, Kunio
为了折叠非天然蛋白质，伴侣蛋白GroEL在ATP依赖性反应循环中经历许多构象变化和GroES结合。我们使用新开发的快速扫描原子力显微镜构建了高分辨率的实时三维观察系统。使用该系统，我们可视化了GroES与单个GroEL的结合和解离，其寿命为6 s（k = 0.17 s（-1））。我们还捕捉到了ATP/ADP诱导的开-闭构象变化的个人GroEL在qGroES和底物蛋白的情况下。也就是说，ATP/ADP结合的GroEL可以改变其构象“从封闭到开放”，而无需额外的ATP水解。此外，在ADP存在下的开放构象的寿命（类似于1.0 s）明显低于ATP和ATP类似物的寿命（2-3 s），这意味着ADP结合的开放形式在结构上不如ATP结合的开放形式稳定。这些结果表明，GroEL在核苷酸存在下至少有两种不同的开放构象：ATP结合的预水解开放构象和ADP结合的开放构象，开放形式的ATP水解使其开放构象不稳定，并诱导GroEL的“从开放到闭合”的构象变化。
In order to fold non-native proteins, chaperonin GroEL undergoes numerous conformational changes and GroES binding in the ATP-dependent reaction cycle. We constructed the real-time three-dimensional-observation system at high resolution using a newly developed fast-scanning atomic force microscope. Using this system, we visualized the GroES binding to and dissociation from individual GroEL with a lifetime of 6 s (k = 0.17 s(-1)). We also caught ATP/ADP-induced open-closed conformational changes of individual GroEL in the absence of qGroES and substrate proteins. Namely, the ATP/ADP-bound GroEL can change its conformation `from closed to open' without additional ATP hydrolysis. Furthermore, the lifetime of open conformation in the presence of ADP (similar to 1.0 s) was apparently lower than those of ATP and ATP-analogs (2-3 s), meaning that ADP-bound open-form is structurally less stable than ATP-bound open-form. These results indicate that GroEL has at least two distinct open-conformations in the presence of nucleotide; ATP-bound prehydrolysis open-form and ADP-bound open-form, and the ATP hydrolysis in open-form destabilizes its open-conformation and induces the `from open to closed' conformational change of GroEL.