| Autor: |
Takanobu A. Katoh, Takashi Daiho, Kazuo Yamasaki, Stefania Danko, Shoko Fujimura, Hiroshi Suzuki |
| Jazyk: |
angličtina |
| Rok vydání: |
2021 |
| Předmět: |
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| Zdroj: |
Scientific Reports, Vol 11, Iss 1, Pp 1-12 (2021) |
| Druh dokumentu: |
article |
| ISSN: |
2045-2322 |
| DOI: |
10.1038/s41598-021-92986-3 |
| Popis: |
Abstract The sarcoendoplasmic reticulum Ca2+-ATPase (SERCA) transports Ca2+ ions across the membrane coupled with ATP hydrolysis. Crystal structures of ligand-stabilized molecules indicate that the movement of actuator (A) domain plays a crucial role in Ca2+ translocation. However, the actual structural movements during the transitions between intermediates remain uncertain, in particular, the structure of E2PCa2 has not been solved. Here, the angle of the A-domain was measured by defocused orientation imaging using isotropic total internal reflection fluorescence microscopy. A single SERCA1a molecule, labeled with fluorophore ReAsH on the A-domain in fixed orientation, was embedded in a nanodisc, and stabilized on Ni–NTA glass. Activation with ATP and Ca2+ caused angle changes of the fluorophore and therefore the A-domain, motions lost by inhibitor, thapsigargin. Our high-speed set-up captured the motion during EP isomerization, and suggests that the A-domain rapidly rotates back and forth from an E1PCa2 position to a position close to the E2P state. This is the first report of the detection in the movement of the A-domain as an angle change. Our method provides a powerful tool to investigate the conformational change of a membrane protein in real-time. |
| Databáze: |
Directory of Open Access Journals |
| Externí odkaz: |
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