Off-axis rotor in Enterococcus hirae V-ATPase visualized by Zernike phase plate single-particle cryo-electron microscopy. Issue 1 (December 2018)

• Record Type: Journal Article
• Authors: Tsunoda, Jun; Song, Chihong; Imai, Fabiana; Takagi, Junichi; Ueno, Hiroshi; Murata, Takeshi; Iino, Ryota; Murata, Kazuyoshi
• Is Part Of: Scientific reports. Volume 8:Issue 1(2018)
• Page Start: 1
• Page End: 10
• Publication Date: 2018-12
• Subjects: Natural history -- Research -- Periodicals; Biology -- Research -- Periodicals; Physical sciences -- Research -- Periodicals; Earth sciences -- Research -- Periodicals; Environmental sciences -- Research -- Periodicals; 502.85
• DOI: 10.1038/s41598-018-33977-9
• Languages: English
• ISSNs: 2045-2322

Abstract

Abstract Eh V-ATPase is an ATP-driven Na+ pump in the eubacteriaEnterococcus hirae (Eh ). Here, we present the first entire structure of detergent-solubilizedEh V-ATPase by single-particle cryo-electron microscopy (cryo-EM) using Zernike phase plate. The cryo-EM map dominantly showed one of three catalytic conformations in this rotary enzyme. To further stabilize the originally heterogeneous structure caused by the ATP hydrolysis states of the V1 -ATPases, a peptide epitope tag system was adopted, in which the inserted peptide epitope sequence interfered with rotation of the central rotor by binding the Fab. As a result, the map unexpectedly showed another catalytic conformation ofEh V-ATPase. Interestingly, these two conformations identified with and without Fab conversely coincided with those of the minor state 2 and the major state 1 ofThermus thermophilus V/A-ATPase, respectively. The most prominent feature inEh V-ATPase was the off-axis rotor, where the cytoplasmic V1 domain was connected to the transmembrane Vo domain through the off-axis central rotor. Furthermore, compared to the structure of ATP synthases, the larger size of the interface between the transmembrane a-subunit and c-ring ofEh V-ATPase would be more advantageous for active ion pumping.