Small molecule activates citrullination through targeting PAD2

Abstract

Citrullination is a post-translational modification catalysed by peptidyl arginine deiminase (PAD) enzymes, and dysregulation of protein citrullination is involved in various pathological disorders. During the past decade, a panel of citrullination inhibitors has been developed, while small molecules activating citrullination have rarely been reported so far. In this study, we screened citrullination activator using an antibody against citrullinated histone H3 (cit-H3), and a natural compound demethoxycurcumin (DMC) significantly activated citrullination. The requirement of PAD2 for DMC-activated citrullination was confirmed by a loss of function assay. Notably, DMC directly engaged with PAD2, and showed binding selectivity among PAD family enzymes. Point mutation assay indicated that residue E352 is essential for DMC targeting PAD2. Consistently, DMC induced typical phenotypes of cells with dysregulation of PAD2 activity, including citrullination-associated cell apoptosis and DNA damage. Overall, our study not only presents a strategy for rationally screening citrullination activators, but also provides a chemical approach for activating protein citrullination. This article is part of the Theo Murphy meeting issue ‘The virtues and vices of protein citrullination’.

A cyclic peptide toolkit reveals mechanistic principles of peptidylarginine deiminase IV (PADI4) regulation

“…Thus, it is conceivable that, by binding in this same region, PADI4_11 promotes this same rearrangement allowing the catalytically-competent conformation of PADI4 to form at reduced calcium concentrations, either by increasing the calcium binding affinity for one of the binding sites or by removing the requirement for calcium binding in one of these sites. It is worth noting that molecular docking studies suggest that the PADI2 activator DMZ potentiates PADI2 activity by binding at a similar site, though whether it reduces the requirement of PADI2 for calcium is unknown 37 . Darrah et al ., also previously reported a class of anti-PADI4 autoantigens from rheumatoid arthritis patients that decreased the concentration of calcium required for PADI4 activity 38 .…”

“…This may enable rational design of PADI inhibitors that block activation, rather than competing for substrate binding, and which might consequently achieve greater isozyme selectivity by binding to a region of the proteins that is less conserved than the active site. The anti-parasitic and anti-viral compound nitazoxanide (NTZ) and the natural product demethoxycurcumin (DMC) have recently been identified as activators of PADI2 36,37 . NTZ increases PADI2-dependent cellular citrullination levels through stabilising PADI2 and thereby increasing total enzyme levels.…”

Citrullination: new tricks for an old mod

“…In their article, Barasa & Thompson discuss the currently available isozyme-specific PADI inhibitors and their properties. Additionally, a study by Ke and colleagues reports a naturally occurring PADI2 activating molecule, demethoxycurcumin (Zhang et al [ 29 ]). Along with nitazoxanide, which was previously described by the same team [ 30 ], this compound may be of significant value in cancer biology contexts where PADI2 is deregulated [ 30 ].…”