A highly selective Hsp90 affinity chromatography resin with a cleavable linker

Area of Science:

• Biochemistry
• Proteomics
• Cancer Biology

Background:

• Heat shock protein 90 (Hsp90) is a therapeutic target in cancer, interacting with over 200 proteins.
• Identifying Hsp90 clients is difficult due to low-affinity, transient interactions.
• Existing methods struggle to selectively isolate native Hsp90 complexes.

Purpose of the Study:

• To develop a novel chemo-proteomic strategy for capturing native Hsp90-client complexes.
• To create a selective Hsp90 affinity resin for identifying physiologically relevant Hsp90 targets.
• To investigate the impact of linker choice on affinity media selectivity.

Main Methods:

• Development of a novel, orthogonally cleavable Hsp90 affinity resin.
• Utilizing a chemo-proteomic strategy for protein purification.
• Testing resin selectivity under low stringency conditions.

Main Results:

• The novel resin selectively purifies native Hsp90 over related proteins (GRP94, TRAP1).
• The resin enables rapid and unambiguous capture of native Hsp90 in complex with native clients.
• Linker choice significantly influences the selectivity of the affinity media.

Conclusions:

• The developed Hsp90 affinity resin is a powerful tool for identifying native Hsp90 clients.
• This method overcomes challenges associated with transient and low-affinity interactions.
• The findings provide a new approach for understanding Hsp90's role in cancer and other diseases.