Unveiling the Role of Hidden Isomers in Large Stokes Shift in mKeima: Harnessing pH-Sensitive Dual-Emission in Bioimaging

Abstract

Elucidating the origin of large Stokes shift (LSS) in certain fluorescent proteins absorbing in blue/blue-green and emitting in red/far-red has been quite illusive. Using a combination of spectroscopic measurements, corroborated by theoretical calculations, the presence of four distinct forms of the chromophore of the red fluorescent protein mKeima is confirmed, two of which are found to be emissive: a feeble bluish-green fluorescence (∼520 nm), which is enhanced appreciably in a low pH or deuterated medium but significantly at cryogenic temperatures, and a strong emission in red (∼615 nm). Using femtosecond transient absorption spectroscopy, the trans-protonated form is found to isomerize within hundreds of femtoseconds to the cis-protonated form, which further yields the cis-deprotonated form within picoseconds followed by structural reorganization of the local environment of the chromophore. Thus, the mechanism of LSS is substantiated to proceed via stepwise excited-state isomerization followed by proton transfer involving three isomers, leaving the fourth one (trans-deprotonated) as a bystander. The exquisite pH sensitivity of the dual emission is further exploited in fluorescence microscopy.

BODIPY-based regioisomers and a donor–acceptor rotor as organic photosensitizers for maximizing singlet oxygen quantum yields and for the photooxidation of thioanisole

“…Details of the experimental setup can be found elsewhere. 36,37 A pump pulse centered at 400 nm was chosen to photoexcite the sample, followed by a broadband white light probe pulse with a varying time delay. This delay allows us to measure the pump-induced changes in absorbance (ΔOD) in the visible wavelength range, providing information about the properties of the excited state.…”

Boosting photocatalytic efficiency of iridium polypyridyl complex in coupling reaction through the induction of ferrocene: insights into bimetallic catalysis

“…6E) based on four exponential components. A comprehensive description of the TA setup and data analysis procedure is discussed in section 14 in ESI † 17 . Resulting evolution-associated decay spectra (EADS) and corresponding population dynamics are presented in Fig.…”

Dual Photoreactive Ternary Ruthenium(II) Terpyridyl Complexes: A Comparative Study on Visible-Light-Induced Single-Step Dissociation of Bidentate Ligands and Generation of Singlet Oxygen

“…We utilized femtosecond transient absorption spectroscopy (fs-TAS) to gain unprecedented insights into the excited-state dynamics of complexes [ 1 ]–[ 4 ] in an aerated medium. The details of the experimental setup can be found elsewhere. , The 2D contour plots of TA data are shown in Figure a,d, and the spectral traces at different pump–probe delays are shown in Figure b,e for complexes [ 1 ] and [ 2 ] in DMSO, respectively. The excitation of [ 1 ] in DMSO into the higher energy side of the 1 MLCT transition (λ ex = 400 nm) is expected to lead to the population of the Ru (dπ) → ptpy (π*) 1 MLCT excited state, resulting in the fs-TAS contour plots, as shown in Figure a.…”