Development of iodonium based gram negative antibacterial compounds and target identification using a chemoproteomics approach

Abstract

Antimicrobial resistance AMR especially among Gram negative pathogens is a major problem worldwide and new classes of chemotherapeutics with novel mechanisms are urgently needed Diphenyleneiodonium DPIC chloride is an antiseptic agent with excellent activity against Gram positive bacteria and moderate inhibitory activity against Gram negative bacteria A number of new approaches towards developing therapeutics against Gram negative pathogens are in urgent need Although several mechanistic studies have been conducted primarily in mammalian cells the biological targets for DPIC remain to be elucidated in bacteria In order to understand the mechanism of action an iodonium probe containing an alkyne handle was next synthesized This probe enables biorthogonal reactions and aids in identifying proteins that the probe and by extension DPI has modified Preliminary findings using activity based protein profiling ABPP methods with bacterial lysate revealed that the iodonium compound modifies proteins with preliminary data suggesting that this modification was covalent and through a cysteine residue Next in order to find the biological targets of DPIC LC MS MS based competitive ABPP was carried out with E coli This analysis revealed that proteins with redox cofactors as well as those involved in respiration were the most likely biological targets for DPIC In order to validate these results one of the target proteins NuOF was cloned purified and was found to be covalently modified by the probe likely through a cysteine residue Having identified targets of DPI in bacteria we next proceeded to perform a structure activity relationship study with a goal of improving potency against Acinetobacter baumannii A baumannii which is listed among the top priority pathogens for new drug development by the World Health Organization Given the abundance of heterocyclic compounds in bioactive molecules such as antibiotics a series of heterocyclic iodonium analogs were synthesized to enhance the spectrum the antibacterial activity