Investigation into Fo/F1 coupling of the FoF1 ATP synthase by time resolved single particle cryo-EM

2016 Fiscal Year Final Research Report

• Project/Area Number: 15K14464
• Research Category: Grant-in-Aid for Challenging Exploratory Research
• Allocation Type: Multi-year Fund
• Research Field: Structural biochemistry
• Research Institution: University of Hyogo
• Principal Investigator: Gerle Christoph 兵庫県立大学, 生命理学研究科, 特任准教授 (10561970)
• Project Period (FY): 2015-04-01 – 2017-03-31
• Keywords: membrane protein / molecular machine / rotational catalysis / mitochondria / single particle cryo-EM / structural dynamics / sample preparation / protein purification

Outline of Final Research Achievements

Mitochondrial FoF1-ATP synthase is central to energy conversion in all mammalian cells and employs a highly efficient mechanism of rotary catalysis to recycle cellular ATP from ADP and Pi. The structural basis for the efficient coupling between membrane bound Fo domain and matrix F1 domain remains unresolved due to the absence of X-ray crystal structures of the intact enzyme and the difficulty of visualizing large scale movements by X-ray crystallography in general. Here we took up the challenge of using the emerging technique of single particle cryo-EM to tacke this important biological problem. Purification of the bovine enzyme from cow heart muscle tissue was successfully established yielding more than 100 mg of excellent sample on a reproducible basis. A novel sample preparation approach named GraDeR was developed and successfully applied to monomeric bovine FoF1 ATP synthase and first structures obtained at a resolution of ~11 Angstrom.

Free Research Field

生物学