Related Experiment Video
Updated: Apr 19, 2026
Genetically-encoded Molecular Probes to Study G Protein-coupled Receptors
Published on: September 13, 2013
Regulation of target protein knockdown and labeling using ligand-directed Ru(bpy)3 photocatalyst
Shinichi Sato1, Kohei Morita, Hiroyuki Nakamura
1Chemical Resources Laboratory, Tokyo Institute of Technology , Yokohama 226-8503, Japan.
Abstract:
Ligand-directed Ru(bpy)3 photocatalysts induce chromophore-assisted light inactivation (CALI) of target proteins under visible light irradiation in vitro and within cells. Here, histidine, methionine, and tryptophan residues were oxidized by the singlet oxygen ((1)O2) generated by Ru(bpy)3 with light. The addition of a tyrosyl radical trapper (TRT), such as N'-acyl-N,N-dimethyl phenylenediamine, inhibited peptide/protein oxidation and induced labeling on the tyrosine residue. This mechanistic study suggests that TRT scavenges (1)O2, concomitant with the coupling reaction to the tyrosyl radical generated by Ru(bpy)3. Both CALI and labeling can be regulated by the Ru(bpy)3 photocatalysts in the absence or presence of TRT. Ligand-conjugated Ru(bpy)3 photocatalysts (local environmental single-electron transfer catalysts: LSCs) were used not only for target-selective protein labeling, but also for protein knockdown by CALI.
Insights
Ruthenium (Ru(bpy)3) photocatalysts enable targeted protein inactivation (CALI) and labeling using visible light. A tyrosyl radical trapper (TRT) modifies this process, allowing for specific tyrosine labeling and enhanced control over protein manipulation.
Area of Science:
- Photochemistry
- Biochemistry
- Molecular Biology
Background:
- Visible light-activated photocatalysts offer precise control over biological processes.
- Chromophore-assisted light inactivation (CALI) is a method for targeted protein function disruption.
- Understanding the reactive oxygen species (ROS) involved in photocatalysis is crucial for developing new tools.
Purpose of the Study:
- To elucidate the mechanism of Ru(bpy)3-mediated CALI and protein labeling.
- To investigate the role of singlet oxygen and tyrosyl radicals in the reaction.
- To explore the application of ligand-conjugated Ru(bpy)3 photocatalysts (LSCs) for protein manipulation.
Main Methods:
- Utilized Ru(bpy)3 photocatalysts with visible light irradiation.
- Investigated the oxidation of histidine, methionine, and tryptophan residues.
- Employed a tyrosyl radical trapper (TRT) to study reaction pathways.
- Analyzed protein labeling and inactivation in vitro and in cellulo.
Main Results:
- Ru(bpy)3 photocatalysts generated singlet oxygen ((1)O2), oxidizing amino acid residues.
- TRT inhibited (1)O2-mediated oxidation and induced tyrosine-specific labeling.
- TRT scavenges (1)O2 and couples with Ru(bpy)3-generated tyrosyl radicals.
- Both CALI and labeling were controllable by Ru(bpy)3 photocatalysts with or without TRT.
Conclusions:
- Mechanistic insights into Ru(bpy)3-mediated photocatalysis were gained.
- TRT provides a means to redirect the photocatalytic reaction towards tyrosine labeling.
- Ligand-conjugated Ru(bpy)3 photocatalysts (LSCs) enable targeted protein labeling and CALI for protein knockdown.
More Related Videos
10:49Identification of Small Molecule-binding Proteins in a Native Cellular Environment by Live-cell Photoaffinity Labeling
Published on: September 20, 2016
09:32Light-mediated Reversible Modulation of the Mitogen-activated Protein Kinase Pathway during Cell Differentiation and Xenopus Embryonic Development
Published on: June 15, 2017