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Title: Mapping the Complete Photocycle that Powers a Large Stokes Shift Red Fluorescent Protein
Abstract

Large Stokes shift (LSS) red fluorescent proteins (RFPs) are highly desirable for bioimaging advances. The RFP mKeima, with coexistingcis‐andtrans‐isomers, holds significance as an archetypal system for LSS emission due to excited‐state proton transfer (ESPT), yet the mechanisms remain elusive. We implemented femtosecond stimulated Raman spectroscopy (FSRS) and various time‐resolved electronic spectroscopies, aided by quantum calculations, to dissect thecis‐ andtrans‐mKeima photocycle from ESPT, isomerization, to ground‐state proton transfer in solution. This work manifests the power of FSRS with global analysis to resolve Raman fingerprints of intermediate states. Importantly, the deprotonatedtrans‐isomer governs LSS emission at 620 nm, while the deprotonatedcis‐isomer's 520 nm emission is weak due to an ultrafastcis‐to‐transisomerization. Complementary spectroscopic techniques as a table‐top toolset are thus essential to study photochemistry in physiological environments.

 
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NSF-PAR ID:
10387325
Author(s) / Creator(s):
 ;  ;  ;  ;  ;  ;  ;  ;  ;  
Publisher / Repository:
Wiley Blackwell (John Wiley & Sons)
Date Published:
Journal Name:
Angewandte Chemie
Volume:
135
Issue:
5
ISSN:
0044-8249
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
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