冷冻电镜研究野生型DegP降解底物的构象变化
秦逸澄，刘蕴辉, 沈庆涛*
（1.上海科技大学生命科学学院，上海201210；2.青岛海洋科学与技术国家实验室，山东青岛266005；3.南方科技大学生命科学学院，广东深圳518005）
摘 要 研究生物大分子结构的动态变化是结构生物学未来发展的重要方向之一，冷冻电镜的三维结构分类技术为实现这一目标提供了可能。本研究以大肠杆菌蛋白酶DegP为研究对象，利用冷冻电镜三维分类技术解析了低温和热休克温度下野生型DegP降解底物的不同构象，比较了不同温度下野生型DegP寡聚状态的变化，指出了相较于酶活丧失突变体，野生型DegP包裹底物后具有更高的结构柔性。该研究成果为热休克条件下DegP通过寡聚降解底物提供了直接证据。
关键词 高温冷冻电镜；DegP笼状结构；动态过程
中图分类号：Q7；Q71；Q336 文献标识码：A doi:10.3969/j.issn.1000-6281.2023.01.005
Cryo-EM reveals cage assembly flexibility of wild-type DegP
QIN Yi-cheng1，LIU Yun-hui 2, SHEN Qing-Tao1，3*
（1.School of Life Science and Technology, ShanghaiTech University，Shanghai 201210；2.Qingdao National Laboratory for Marine Science and Technology, Qingdao Shandong 266237；3. School of Life Science, South University of Science and Technology，Shenzhen Guangdong 518005，China）
Abstract 3D classification and the derived variability analysis in cryo-electron microscopy offer great opportunity to investigate the structural dynamics of biological macromolecules. Here, we utilized wild-type DegP as a model system and investigated its cage assembly in the presence of denatured substrate under low or high temperatures. Our 3D reconstructions reveal cage assembly difference of wild-type DegP under different temperatures and a more flexible cage relative to protease-deficient mutant. Our results provide direct evidence that cage assembly is required for proteolytic activity of DegP and demonstrate the important role of Cryo-EM in the protein dynamics study.
Keywords high-temperature Cryo-EM；DegP cage assembly；dynamic structures
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