Abstract
Structure-based drug design is an emerging technology for developing new drugs. However, in silico modeling and predicting inhibitors covalently bound to cofactor-containing enzymes remain challenging. Here, we demonstrate an alternative approach for visualizing protein inhibitor binding via X-ray crystallography of PLP-dependent enzyme crystals. Specifically, we used crystals of type II cysteine desulfurase, SufS, a drug-target enzyme in the iron-sulfur cluster biosynthetic SUF system. We identified (2R,3R)-3-ethoxycarbonylaziridine-2-carboxylic acid (EAC) as a selective inhibitor of SufS. The X-ray crystal structures of SufS soaked with EAC for 12 and 24 h showed PLP-ligand conjugates. Two PLP-ligand conjugate species were assigned to l-α-formylglycine-PLP external aldimine and aminomalonate-PLP external aldimine, which could be caused by the aziridine ring opening reaction, removal of the ethyl ester, and air oxidation. This strategy could help identify new drug candidates specific to SufS, a new drug target in pathogenic microorganisms containing iron-sulfur cluster biosynthetic SUF systems.