Structural information about the trans-to-cis isomerization mechanism of the photoswitchable fluorescent protein rsEGFP2 revealed by multiscale infrared transient absorption

Uriarte, Lucas, Vitalle, Rafaelle, Nizinski, Stanizlaw, Hadjidemetriou, Kyprianos, Zala, Ninon, Lukacs, Andras, Greetham, Gregory M., Sazanovich, Igor, Weik, Martin, Ruckebusch, Cyril, Meech, Steve and Sliwa, Michel (2022) Structural information about the trans-to-cis isomerization mechanism of the photoswitchable fluorescent protein rsEGFP2 revealed by multiscale infrared transient absorption. The Journal of Physical Chemistry Letters, 13 (5). 1194–1202. ISSN 1948-7185

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Abstract

RsEGFP2 is a reversibly photoswitchable fluorescent protein used in super-resolved optical microscopies, which can be toggled between a fluorescent On state and a nonfluorescent Off state. Previous time-resolved ultraviolet–visible spectroscopic studies have shown that the Off-to-On photoactivation extends over the femto- to millisecond time scale and involves two picosecond lifetime excited states and four ground state intermediates, reflecting a trans-to-cis excited state isomerization, a millisecond deprotonation, and protein structural reorganizations. Femto- to millisecond time-resolved multiple-probe infrared spectroscopy (TRMPS-IR) can reveal structural aspects of intermediate species. Here we apply TRMPS-IR to rsEGFP2 and implement a Savitzky–Golay derivative analysis to correct for baseline drift. The results reveal that a subpicosecond twisted excited state precursor controls the trans-to-cis isomerization and the chromophore reaches its final position in the protein pocket within 100 ps. A new step with a time constant of 42 ns is reported and assigned to structural relaxation of the protein that occurs prior to the deprotonation of the chromophore on the millisecond time scale.

Item Type: Article
Uncontrolled Keywords: materials science(all),physical and theoretical chemistry ,/dk/atira/pure/subjectarea/asjc/2500
Faculty \ School: Faculty of Science > School of Chemistry
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Depositing User: LivePure Connector
Date Deposited: 22 Apr 2022 09:30
Last Modified: 28 Apr 2022 08:31
URI: https://ueaeprints.uea.ac.uk/id/eprint/84733
DOI: 10.1021/acs.jpclett.1c02920

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