Fig. 1. Structure-based design leads to an electrophilic PARP16 inhibitor that covalently modifies a non-conserved cysteine. (A) Crystal structure of the active site of PARP16 (orange, PDB: 4F0D) overlaid with PARP1 (cyan, PDB: 5DS3) bound to olaparib (green). (B) PARP family sequence alignment generated with T-Coffee multiple sequence alignment algorithm. The non-conserved D-loop cysteine (Cys169) of PARP16 is highlighted in red. (C) Structure of HJ-52 and DB008, with the acrylamide selectivity element shown in red, and the dual selectivity element/clickable alkyne handle shown in green. Biochemical activity assay to assess potency of olaparib, HJ-52, and DB008 against PARP16 (D) and PARP1 (E); n ≥ 3 biological replicates.