Lighting Up the Plasma Membrane: Development and Applications of Fluorescent Ligands for Transmembrane Proteins

Abstract

Despite the fact that transmembrane proteins represent the main therapeutic targets for decades, complete and in‐depth knowledge about their biochemical and pharmacological profiling is not fully available. In this regard, target‐tailored small‐molecule fluorescent ligands are a viable approach to fill in the missing pieces of the puzzle. Such tools, coupled with the ability of high‐precision optical techniques to image with an unprecedented resolution at a single‐molecule level, helped unraveling many of the conundrums related to plasma proteins’ life‐cycle and druggability. Herein, we review the recent progress made during the last two decades in fluorescent ligand design and potential applications in fluorescence microscopy of voltage‐gated ion channels, ligand‐gated ion channels and G‐coupled protein receptors.

A versatile Halo- and SNAP-tagged BMP/TGFβ receptor library for quantification of cell surface ligand binding

“…Interrogating cell surface receptor biology in living cells, including their distribution, stoichiometries, ligand-binding efficacies and downstream activation has been pushed in the last years mainly by state-of-the-art microscopy techniques (Borgarelli et al, 2021; Kemter et al, 2021). In all these instances, the choice of labelling the protein of interest is of immense importance (Sahl, Hell, & Jakobs, 2017).…”

Fluorescent Benzothiadiazole Derivatives as Fluorescence Imaging Dyes: A Decade of New Generation Probes

“…Because of the importance of the PM to life maintenance, small fluorescent organic dyes have been used to investigate the PM and the specificities associate with this complex and dynamic structure. [199] Several fluorescent organic molecules were recently developed to visualize PM selectively and to depict the cellular events associate with this essential structure. [200][201][202][203][204][205][206][207][208][209] Although the importance of the PM, only very recently BTD based bioprobes have been reported exclusively by our group as selective fluorescence cell imaging probes (Figure 24).…”

“…Because of the importance of the PM to life maintenance, small fluorescent organic dyes have been used to investigate the PM and the specificities associate with this complex and dynamic structure [199] . Several fluorescent organic molecules were recently developed to visualize PM selectively and to depict the cellular events associate with this essential structure [200–209] .…”

N-Cyanorhodamines: cell-permeant, photostable and bathochromically shifted analogues of fluoresceins

“…Thus, fluoresceins or negatively charged sulfonated rhodamine labels are often preferred in this context. 23 Moreover, the use of positively charged rhodamine probes in live-cell imaging is severely limited by their off-targeting to mitochondria. 24 Unlike fluoresceins, rhodamine labels remain fluorescent in acidic media and will appear as bright granules whenever accumulated in lysosomes (pH 4.6–5.0) and endosomes (pH 5–6), 9 which prompted the development of more complex self-quenching fluorogenic rhodamine–tetrazine conjugates 25 for wash-free imaging.…”