2016 Fiscal Year Annual Research Report
Investigation into Fo/F1 coupling of the FoF1 ATP synthase by time resolved single particle cryo-EM
| Project/Area Number |
15K14464
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| Research Institution | University of Hyogo |
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Principal Investigator |
Gerle Christoph 兵庫県立大学, 生命理学研究科, 特任准教授 (10561970)
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| Project Period (FY) |
2015-04-01 – 2017-03-31
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| Keywords | membrane protein / molecular machine / rotational catalysis / mitochondria / cryo-EM / single particle / structural dynamics / sample preparation |
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| Outline of Annual Research Achievements |
In the spirit of the Hoga Kakenhi grant this project aims high at using single particle cryo-EM to obtain a time resolved structural analysis of the mammalian F-ATP synthase during rotary catalysis. Structural dynamics of rotary catalysis is an an important scientific problem. It is difficult to tackle by X-ray crystallography and ways and means to solve this problem will be influential for improving the understanding of other membrane bound molecular machines. We have succeeded in establishing a mild purification procedure and importantly innovated a new sample preparation method for the use of cryo-EM of fragile membrane complexes termed GraDeR. We successfully applied GraDeR to the cryo-EM analysis of mammalian F-ATP synthase and obtained new insights into the structure of this important mitochondrial energy converter. Furthermore, we were able to apply focused masking to cryoEM image data sets to resolve critical dynamics in the transmembrane Fo domain. Taken together, the foundations of using time resolved cryoEM for analyzing the structural dynamics of mammalian F-ATP synthase were firmly established and thus the door for a whole new series of cryo-EM studies on mammalian F-ATP synthase opened.
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| Remarks |
The pdf of the step-by-step protocol has been already downloaded several hundred times and the GraDeR web-page is well known in the community.
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[Journal Article] Two-dimensional crystallization of monomeric bovine cytochrome c oxidase with bound cytochrome c in reconstituted lipid membranes.2016
Author(s)
Osuda, Y., Shinzawa-Itoh, K., Tani, K., Maeda, S., Yoshikawa, S., Tsukihara, T. and Gerle C.
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Journal Title
Microscopy
Volume: 65
Pages: 263-267
DOI
Peer Reviewed / Open Access / Int'l Joint Research
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[Presentation] GraDeR:単粒子解析等の膜タンハク質資料調整2016
Author(s)
Christoph Gerle, Florian Hauer, Niels Fischer, Kyoko Shinzawa-Itoh, Satoru Shimada, Ken Yokoyama, Atsunori Oshima, Yoshinori Fujiyoshi, Holger Stark
Organizer
日本生物物理学会
Place of Presentation
つくば国際会議場（茨城県、つくば市）
Year and Date
2016-11-25 – 2016-11-27
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[Presentation] Electron crystallography of tomographic volumes.2016
Author(s)
Kazutoshi Tani, Karen Davies, Chimari Jiko, Shintaro Maeda, Kyoko Shinzawa-Itoh, Deryck Mills, Tomitake Tsukihara, Yoshinori Fujiyoshi, Werner Kuhlbrandt, Christoph Gerle
Organizer
3DEM Gordon Research Conference
Place of Presentation
Hong Kong University, Hong Kong
Year and Date
2016-06-19 – 2016-06-24
Int'l Joint Research
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