A Maleimide‐functionalized Tetraphenylethene for Measuring and Imaging Unfolded Proteins in Cells

Abstract

Collapse of the protein homeostasis (proteostasis) can lead to accumulation and aggregation of unfolded proteins, which has been found to associate with a number of disease conditions including neurodegenerative diseases, diabetes and inflammation. Here we report a maleimide‐functionalized tetraphenylethene (TPE)‐derivatized fluorescent dye, TPE‐NMI, which shows fluorescence turn‐on property upon reacting with unfolded proteins in vitro and in live cells under proteostatic stress conditions. The level of unfolded proteins can be measured by flow cytometry and visualized with confocal microscopy.

Rational Design of Crystallization‐Induced‐Emission Probes To Detect Amorphous Protein Aggregation in Live Cells

“…Another challenge for selective detection of amorphous aggregates is how to achieve selectivity over other organelles in the complex cellular milieu as they accumulate intracellularly rather than the extracellular space for amyloid aggregates. Although AIEgens exhibit appealing performance in detecting amyloid aggregated proteins, they failed to detect cellular amorphous ones without the covalent linkage to proteins exemplified by the seminal works from Hong and Zhang et al [34–38] . Sensors of inherent selective binding towards amorphous aggregated proteins, however, were seldom reported.…”

A Molecular Chameleon for Mapping Subcellular Polarity in an Unfolded Proteome Environment

“…To achieve the environmental sensitivity, our strategy involved replacing the TPE unit with a push‐pull AIE fluorophore (Figure A). Push‐pull dyes with both electron donor and acceptor experience excited‐state charge transfer, which is stabilized by interaction with the dipoles of solvent, resulting in emission redshift in more polar solvents . Being sensitive to polarity, push‐pull dyes have been extensively used to study the biophysical properties of lipid membranes and for the detection of biomolecular interactions such as protein–lipid interactions .…”

“…To construct a push‐pull AIE fluorophore while retaining a similar molecular structure, we changed one of the phenyl rings of TPE into an electron‐withdrawing cyano group (Figure A) . Methoxy and dimethylamino substituents are used as electron donors, which also improve the compatibility of the dyes in aqueous media …”

“…In addition, we observed that electron density for the HOMO levels was more localized around the electron‐donating groups while that for the LUMO was closer to the electron‐withdrawing cyano group (Figure B). Such electron distribution is characteristic of push‐pull dyes with photoinduced charge transfer, which results in the solvatochromism in their fluorescence . To study the fluorescence behavior, the thiol‐bound form of NTPAN‐MI ( 4 a ) was prepared (Figure D; see Scheme S1).…”

“…We further apply our strategy to quantify the change of polarity surrounding unfolded proteins in different intracellular compartments driven by the increase of the unfolded protein load. Compared to our previously reported approaches to quantifying cytoplasmic unfolded proteins, this multiresponsive probe serves as a “molecular chameleon” that holds unprecedented advantage for direct visualizing and quantifying unfolded proteins in both the cytoplasm and the nucleus, with sensitivity to in situ environmental polarity changes in different subcellular compartments arising from protein unfolding.…”