A FluoPol-ABPP PAD2 high-throughput screen identifies the first calcium site inhibitor targeting the PADs.
The protein arginine deiminases (PADs) catalyze the post-translational hydrolysis of peptidyl-arginine to form peptidyl-citrulline in a process termed deimination or citrullination. PADs likely play a role in the progression of a range of disease states because dysregulated PAD activity is observed in a host of inflammatory diseases and cancer. For example, recent studies have shown that PAD2 activates ERα target gene expression in breast cancer cells by citrullinating histone H3 at ER target promoters. To date, all known PAD inhibitors bind directly to the enzyme active site. PADs, however, also require calcium ions to drive a conformational change between the inactive apo-state and the fully active calcium bound holoenzyme, suggesting that it would be possible to identify inhibitors that bind the apoenzyme and prevent this conformational change. As such, we set out to develop a screen that can identify PAD2 inhibitors that bind to either the apo or calcium bound form of PAD2. Herein, we provide definitive proof of concept for this approach and report the first PAD inhibitor, ruthenium red (Ki of 17 μM), to preferentially bind the apoenzyme.
Department of Chemistry, ‡Lead ID, and ∥The Kellogg School of Science and Technology, The Scripps Research Institute , Scripps Florida, 120 Scripps Way, Jupiter, Florida 33458, United States.
Daniel M
DM
Lewallen
Kevin L
KL
Bicker
Franck
F
Madoux
Peter
P
Chase
Lynne
L
Anguish
Scott
S
Coonrod
Peter
P
Hodder
Paul R
PR
Thompson
D001665:Binding Sites; D001681:Biological Assay; D002118:Calcium / Q000737:chemistry*; D016503:Drug Delivery Systems; D004353:Drug Evaluation, Preclinical; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006867:Hydrolases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D015394:Molecular Structure; D011485:Protein Binding / Q000187:drug effects; D000076342:Protein-Arginine Deiminases; D012430:Ruthenium Red / Q000737:chemistry* / Q000494:pharmacology*
D004791:Enzyme Inhibitors; D012430:Ruthenium Red; D006867:Hydrolases; D000076342:Protein-Arginine Deiminases; D002118:Calcium