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Single molecule kinetics of bacteriorhodopsin by HS-AFM.

TitleSingle molecule kinetics of bacteriorhodopsin by HS-AFM.
Publication TypeJournal Article
Year of Publication2021
AuthorsPerrino AP, Miyagi A, Scheuring S
JournalNat Commun
Volume12
Issue1
Pagination7225
Date Published2021 12 10
ISSN2041-1723
Abstract

Bacteriorhodopsin is a seven-helix light-driven proton-pump that was structurally and functionally extensively studied. Despite a wealth of data, the single molecule kinetics of the reaction cycle remain unknown. Here, we use high-speed atomic force microscopy methods to characterize the single molecule kinetics of wild-type bR exposed to continuous light and short pulses. Monitoring bR conformational changes with millisecond temporal resolution, we determine that the cytoplasmic gate opens 2.9 ms after photon absorption, and stays open for proton capture for 13.2 ms. Surprisingly, a previously active protomer cannot be reactivated for another 37.6 ms, even under excess continuous light, giving a single molecule reaction cycle of ~20 s-1. The reaction cycle slows at low light where the closed state is prolonged, and at basic or acidic pH where the open state is extended.

DOI10.1038/s41467-021-27580-2
Alternate JournalNat Commun
PubMed ID34893646
Grant ListR01 NS110790 / NS / NINDS NIH HHS / United States
DP1 AT010874 / AT / NCCIH NIH HHS / United States