A “Clickable” Photoconvertible Small Fluorescent Molecule as a Minimalist Probe for Tracking Individual Biomolecule Complexes

Abstract

Photoconvertible fluorophores can enable the visualization and tracking of a specific biomolecules, complexes, and cellular compartments with precise spatiotemporal control. The field of photoconvertible probes is dominated by fluorescent protein variants, which can introduce perturbations to the target biomolecules due to their large size. Here, we present a photoconvertible small molecule, termed CPX, that can be conjugated to any target through azidealkyne cycloaddition ("click" reaction). To demonstrate its utility, we have applied CPX to study 1) trafficking of biologically relevant synthetic vesicles and 2) intracellular processes involved in transmission of α-synuclein (αS) pathology. Our results demonstrate that CPX can serve as a minimally perturbing probe for tracking the dynamics of biomolecules.

Genetic encoding of a highly photostable, long lifetime fluorescent amino acid for imaging in mammalian cells

“…Phe94 and Glu114 were chosen as the incorporation sites as they have been shown to tolerate a variety of bulky uorophore substitutions, including both ncAA incorporation and Cys modication. 16,[43][44][45][46][47][48][49][50] As a positive control, we expressed the same protein construct using an Mj RS previously optimized for Acd incorporation in E. coli (Mj AcdRS A9). 14 As an additional comparison of anticipated masses and yields, we also expressed the wild-type (WT) aS sequence fused to the His 6 intein tag.…”

Achieving highly efficient aggregation-induced emission, reversible and irreversible photochromism by heavy halogen-regulated photophysics and D–A molecular pattern-controlled photochemistry of through-space conjugated luminogens

“…These observations demonstrate that the introduction of dimethylamino groups into the TPAN skeleton not only constructs a unique D-p-A pattern and red-shied emissions but also greatly turns their photochemical behaviours from unstable and reversible cyclization to UV-dependent stable and irreversible cyclization and dehydrogenation. 57,58 Photoswitchable reversible and irreversible patterning on various surfaces and multiple information encryption Most diarylethene-based photochromic molecules with uorescence turn-on behaviors were examined in their solution states, and a few of them exhibited photoswitchable patterning on the solid surface upon irradiation. [26][27][28][29] Herein, we demonstrate that TPAN compounds with reversible photochromism possess the ability to undergo photo-triggered reactions and the resulting sharp color changes in the diverse matrix and on various surfaces.…”

Light-Emitting Probes for Labeling Peptides

“… 35 Also, a water-soluble, cell-permeable, and non-cytotoxic photoconvertible small fluorescent probe called CPX was synthesized ( Figure 2B ; Table 1 ). 36 CPX was coupled at three different positions within the α-synuclein protein through Huisgen 1,3-dipolar azide-alkyne cycloaddition and, due to its small size, overcame the common limitations of FPs in studies involving α-synuclein aggregate dynamics ( Figure 3 ). 36 Moreover, the precursor of this probe can be conjugated to other biomolecules through nucleophilic displacement of the iodide.…”