Development of Nuclear-Targeted Photoaffinity Probes with Cleavable Disulfide Bonds for Enhancing Selectivity in hPTM...

Area of Science:

• Biochemistry
• Molecular Biology
• Proteomics

Background:

• Histone post-translational modification (hPTM) interactome profiling is crucial for understanding cellular processes.
• Existing cell-penetrating peptides, like cyclic polyarginine (cR10), cause interference in these studies due to high positive charge.
• Nuclear-targeted histone photoaffinity probes are needed for accurate interactome analysis.

Purpose of the Study:

• To develop novel nuclear-targeted histone photoaffinity probes with intracellularly cleavable linkages.
• To improve the selectivity and reduce false-positive signals in hPTM interactome profiling.
• To target specific histone marks, H3K4me3 and H3K14la.

Main Methods:

• Facile synthesis of novel nuclear-targeted histone photoaffinity probes.
• Incorporation of intracellularly cleavable cR10-disulfide linkages.
• In vitro labeling and proteomic assays to evaluate probe performance.

Main Results:

• The novel probes demonstrated significantly improved binding selectivity for reader proteins.
• Drastic reduction in false-positive signals was observed after cR10 cleavage.
• Successful targeting of H3K4me3 and H3K14la histone marks.

Conclusions:

• Intracellularly cleavable histone photoaffinity probes offer enhanced accuracy for hPTM interactome profiling.
• The developed probes overcome the limitations of noncleavable peptides, reducing experimental interference.
• This advancement facilitates more precise investigation of histone modification pathways.