Novel Cyclopropene Probes as Chemical Reporters for Bioorthogonal Metabolic Labeling of Benzoylated...

Area of Science:

• Biochemistry
• Chemical Biology
• Cellular Biology

Background:

• Lysine benzoylation (Kbz) is a critical post-translational modification impacting cellular functions.
• Existing detection methods like mass spectrometry lack dynamic tracking capabilities in living cells.
• Previous chemical probes for acylated modifications have limitations for cellular-level dynamic studies.

Purpose of the Study:

• To develop a novel chemical reporter for the dynamic tracking of lysine benzoylation in live cells.
• To evaluate the efficiency and bio-orthogonality of the new probe for metabolic labeling.

Main Methods:

• Development of a 1-methylcyclopropene chemical reporter for IEDDA reaction with S-tetrazine-BODIPY.
• Evaluation of probe's optical properties, kinetic constants, and bio-orthogonality.
• Validation of metabolic labeling in RAW and HepG2 cells and dynamic detection using a SIRT2 inhibitor.

Main Results:

• A novel benzoic acid probe (probe 2) was identified as highly efficient for lysine benzoylation detection.
• The probe demonstrated comparable kinetic parameters to endogenous benzoylated peptides when acted upon by SIRT2.
• Successful metabolic labeling and dynamic monitoring of lysine benzoylation were achieved in cellular models.

Conclusions:

• The developed chemical probe enables efficient and bio-orthogonal metabolic labeling of lysine benzoylation.
• This strategy allows for dynamic tracking of lysine benzoylation levels in live cells.
• The findings establish a foundation for developing new metabolic labeling tools for dynamic post-translational modification studies.