A simple and effective cleavable linker for chemical proteomics applications

Area of Science:

• Chemical proteomics
• Mass spectrometry (MS) based protein identification

Background:

• Isolation and purification of protein targets are crucial steps before MS identification in chemical proteomics.
• The widely used biotin-streptavidin system suffers from harsh denaturing conditions, leading to the release of non-specifically bound and natively biotinylated proteins.
• Existing cleavable linkers are often inefficient, difficult to synthesize, or expensive, highlighting the need for improved tools.

Purpose of the Study:

• To introduce an efficient, easily synthesized, and inexpensive cleavable linker for chemical proteomics applications.
• To demonstrate the utility of a vicinal diol cleavable linker in activity-based protein profiling and bioorthogonal ligation.
• To reduce background protein identifications compared to standard on-bead digestion methods.

Main Methods:

• Incorporation of a vicinal diol cleavable linker into an activity-based probe.
• Development of a biotin alkyne tag utilizing the cleavable linker for bioorthogonal ligation.
• Application of the developed reagents in protein isolation and purification prior to MS identification.

Main Results:

• The vicinal diol cleavable linker was successfully incorporated into chemical proteomics reagents.
• The new reagents facilitated protein target isolation with significantly reduced background identifications.
• The developed linker strategy offers an improvement over standard on-bead digestion techniques.

Conclusions:

• The vicinal diol cleavable linker represents a valuable addition to the chemical proteomics toolbox.
• This linker strategy effectively minimizes non-specific binding and improves the accuracy of protein identification.
• The developed reagents provide an efficient and cost-effective solution for protein enrichment and analysis.