Transition state mimics are valuable mechanistic probes for structural studies with the arginine methyltransferase CARM1

Abstract

Coactivator associated arginine methyltransferase 1 (CARM1) is a member of the protein arginine methyltransferase (PRMT) family and methylates a range of proteins in eukaryotic cells. Overexpression of CARM1 is implicated in a number of cancers, and it is therefore seen as a potential therapeutic target. Peptide sequences derived from the well-defined CARM1 substrate poly(A)-binding protein 1 (PABP1) were covalently linked to an adenosine moiety as in the AdoMet cofactor to generate transition state mimics. These constructs were found to be potent CARM1 inhibitors and also formed stable complexes with the enzyme. High-resolution crystal structures of CARM1 in complex with these compounds confirm a mode of binding that is indeed reflective of the transition state at the CARM1 active site. Given the transient nature of PRMT–substrate complexes, such transition state mimics represent valuable chemical tools for structural studies aimed at deciphering the regulation of arginine methylation mediated by the family of arginine methyltransferases

Structural and biochemical evaluation of bisubstrate inhibitors of protein arginine N-methyltransferases PRMT1 and CARM1 (PRMT4)

“…In line with recent advances [38], the development and structural investigation of rationally designed bisubstrate inhibitors remains a desirable route to identify individual residue variations that may contribute to selective inhibition of CARM1 over PRMT1 for example. (B) Structures of bisubstrate azo-SAM inhibitors from the present study (4-10) and those reported previously by van Haren et al [28,33].…”

“…In both cases, ternary crystal structures with CARM1-SAH showed the inhibitor to occupy the substrate-binding pocket with kinetic data supporting noncompetitive inhibition with respect to both SAM and peptide substrate [23,32]. Alternatively, inhibitors may occupy both the SAM and arginine binding sites ( Figure 1B) [28,[33][34][35]. A number of such inhibitors that target CARM1 feature amino adenosine linked to modified cytosine moieties that occupy the SAM and arginine binding site, respectively [36].…”

“…PRMT inhibitors reported to date can be broadly categorised into SAM analogues (including the panmethyltransferase inhibitors sinefungin (3) and S-adenosyl homocysteine (2), ( Figure 1A)), substrate peptide analogues [25][26][27][28], and structures optimised from high throughput screening hits [29][30][31][32]. Promising CARM1 inhibitors include EPZ022302/EZM2302 (a 6 nM CARM1 inhibitor with anti-tumour activity in vivo) [23] and TP-064 (IC 50 < 10 nM against CARM1 and >100-fold selective over other methyltransferases) [32].…”

“…Van Haren et al have also explored this approach by 5 0 -hydroxyl group of adenosine with a methylguanidinium group, which resulted in a compound with 100-fold selectivity for CARM1 over PRMT1 [33]. Extending the guanidinium group to a CARM1 substrate peptide achieved a greater degree of selectivity for CARM1 over PRMT1 (IC 50 values of 0.09 and 25.5 mM, respectively for the most selective compound) [28]. Whilst promising in their apparent selectivity, a key drawback of such peptide-fused mimics is their potential susceptibility to proteolysis, which limits their utility in cellular assays and in vivo applications.…”

Hijacking DNA methyltransferase transition state analogues to produce chemical scaffolds for PRMT inhibitors

“…-, not determined. [29,48] (electronic supplementary material, figure S1). For all inhibitors, the interactions with the base and sugar moieties are strictly conserved and identical to what is observed for SAH [46].…”