Chaperonins GroEL and GroES: views from atomic force microscopy.
Chaperonins GroEL and GroES: views from atomic force microscopy.
复制标题
伴侣蛋白 GroEL 和 GroES：来自原子力显微镜的视图。
DOI：
10.1016/s0006-3495(96)79422-5
复制
发表时间：
1996
影响因子：
3.4
通讯作者：
Shao,Z
中科院分区：
文献类型：
--
作者：
Mou,J;Sheng,S;Ho,R;Shao,Z
用原子力显微镜(AFM)对大肠杆菌伴侣蛋白、GroEL和GroES及其在非水解性核苷酸AMP-PNP存在下的复合体进行了成像。我们证明了吸附在云母表面的GroEL和GroES都可以在室温下被AFM直接在水溶液中分解。然而，由于戊二醛固定了已经吸附的分子，GroEL和GroES的分辨率都得到了进一步的提高，因为所有七个亚基都在没有任何图像处理的情况下得到了很好的分辨。我们还发现，化学固定对于接触式AFM成像GroEL/ES复合体以及在AFM图像中是必要的。有Groes绑定的GroEL和没有绑定GroEL的GroEL可以明显区分开来。GroEL/ES复合体比单独的GroEL高约5 nm，表明GroEL的顶端结构域上移了2 nm。使用稍大的探针力，可以解剖未固定的GroEL：移除上层七聚体以暴露两个七聚体的接触面。这些结果清楚地证明了交联剂在原子力显微镜确定分子结构中的作用。它们也为利用原子力显微镜研究GroE系统和其他分子伴侣的功能的结构基础铺平了道路。
The Escherichia coli chaperonins, GroEL and GroES, as well as their complexes in the presence of a nonhydrolyzable nucleotide AMP-PNP, have been imaged with the atomic force microscope (AFM). We demonstrate that both GroEL and GroES that have been adsorbed to a mica surface can be resolved directly by the AFM in aqueous solution at room temperature. However, with glutaraldehyde fixation of already adsorbed molecules, the resolution of both GroEL and GroES was further improved, as all seven subunits were well resolved without any image processing. We also found that chemical fixation was necessary for the contact mode AFM to image GroEL/ES complexes, and in the AFM images. GroEL with GroES bound can be clearly distinguished from those without. The GroEL/ES complex was about 5 nm higher than GroEL alone, indicating a 2 nm upward movement of the apical domains of GroEL. Using a slightly larger probe force, unfixed GroEL could be dissected: the upper heptamer was removed to expose the contact surface of the two heptamers. These results clearly demonstrate the usefulness of cross-linking agents for the determination of molecular structures with the AFM. They also pave the way for using the AFM to study the structural basis for the function of GroE system and other molecular chaperones.