posted on 2024-04-03, 19:33authored byZhonglin Liu, Jarrett R. Remsberg, Haoxin Li, Evert Njomen, Kristen E. DeMeester, Yongfeng Tao, Guoqin Xia, Rachel E. Hayward, Minjin Yoo, Tracey Nguyen, Gabriel M. Simon, Stuart L. Schreiber, Bruno Melillo, Benjamin F. Cravatt
Covalent
chemistry coupled with activity-based protein
profiling
(ABPP) offers a versatile way to discover ligands for proteins in
native biological systems. Here, we describe a set of stereo- and
regiochemically defined spirocycle acrylamides and the analysis of
these electrophilic “stereoprobes” in human cancer cells
by cysteine-directed ABPP. Despite showing attenuated reactivity compared
to structurally related azetidine acrylamide stereoprobes, the spirocycle
acrylamides preferentially liganded specific cysteines on diverse
protein classes. One compound termed ZL-12A promoted the degradation
of the TFIIH helicase ERCC3. Interestingly, ZL-12A reacts with the
same cysteine (C342) in ERCC3 as the natural product triptolide, which
did not lead to ERCC3 degradation but instead causes collateral loss
of RNA polymerases. ZL-12A and triptolide cross-antagonized one another’s
protein degradation profiles. Finally, we provide evidence that the
antihypertension drug spironolactonepreviously found to promote
ERCC3 degradation through an enigmatic mechanismalso reacts
with ERCC3_C342. Our findings thus describe monofunctional degraders
of ERCC3 and highlight how covalent ligands targeting the same cysteine
can produce strikingly different functional outcomes.