Pyridinium Analogues of Green Fluorescent Protein Chromophore: Fluorogenic Dyes with Large Solvent-Dependent Stokes Shift

Abstract

Novel fluorogenic dyes based on the GFP chromophore are developed. The compounds contain a pyridinium ring instead of phenolate and feature large Stokes shifts and solvent-dependent variations in the fluorescence quantum yield. Electronic structure calculations explain the trends in solvatochromic behavior in terms of the increase of the dipole moment upon excited-state relaxation in polar solvents associated with the changes in bonding pattern in the excited state. A unique combination of such optical characteristics and lipophilic properties enables using one of the new dyes for imaging the membrane structure of endoplasmic reticulum. An extremely high photostability (due to a dynamic exchange between the free and absorbed states) and selectivity make this compound a promising label for this type of cellular organelles.

Red‐Shifted Substrates for FAST Fluorogen‐Activating Protein Based on the GFP‐Like Chromophores

“…However, they become highly emissive when the flexible substituent is fixed by an external influence or by an internal lock . Previously they were successfully used as fluorogens for various proteins, nucleic acids, and whole cell organelles …”

“…[12] Previously they weres uccessfully used as fluorogens for various proteins, [13] nucleic acids, [14] and whole cell organelles. [15] Various rhodanine derivatives [16] are close structurala nalogueso fF Pc hromophores ( Figure 1B). Several hydroxybenzylidener hodanines were previously used for fluorescent label- ing in complex with the fluorogen-activating protein, FAST (fluorescence-activating and absorption-shifting tag), [17] an engineeredv ariant of the photoreceptorfrom Halorhodospira halophila,p hotoactive yellow protein (PYP).…”

Fluorescent probes for targeting endoplasmic reticulum: design strategies and their applications

“…74 Finally, the synthetic analogs of HBDI (4 0 -hydroxybenzylidene-2,3-dimethylimidazolinone), GFP chromophore (5q), were synthesized with pyridine replacing the phenyl ring and used to stain the ER efficiently, primarily due to its lipophilic properties. 80…”

“…The membrane-rich ER is characterized by high hydrophobicity, and the favorable hydrophobic interactions with the (5a-5c) led to ER specific localization. 67,80 Three amphiphilic squaraine fluorophores with hydrocarbon chains and zwitterionic polar head (5o) were synthesized by Collot and coworkers. By structural tuning of these molecules, different results of cellular localization were obtained.…”

Regulation of mitochondrial proteostasis by the proton gradient

“…Since the di-aspartyl pH sensor motif of TMBIM6 is conserved in TMBIM5 and since a H + gradient is constantly present across the IM, we examined whether TMBIM5 can act as a Ca 2+ /H + exchanger. We expressed a mitochondrially targeted variant of the synthetic pH sensor SypHer (SypHer3s-dmito, (Ermakova et al, 2018)) in cells depleted of TMBIM5 and monitored the pH in the mitochondrial matrix (Figure 2B). Strikingly, loss of TMBIM5 increased the matrix pH from pH 7.25 to pH 7.6 (Figure 2B), indicating that the TMBIM5-mediated Ca 2+ release from mitochondria is indeed accompanied by H + transport into the mitochondrial matrix.…”