Chemical Proteomics Reveals Antibiotic Targets of Oxadiazolones in MRSA

Abstract

Phenotypic screening is a powerful approach to identify novel antibiotics, but elucidation of the targets responsible for the antimicrobial activity is often challenging in the case of compounds with a polypharmacological mode of action. Here, we show that activity-based protein profiling maps the target interaction landscape of a series of 1,3,4-oxadiazole-3-ones identified in a phenotypic screen to have high antibacterial potency against multidrug-resistant Staphylococcus aureus. In situ competitive and comparative chemical proteomics with a tailor-made activity-based probe, in combination with transposon and resistance studies, revealed several cysteine and serine hydrolases as relevant targets. Our data showcase oxadiazolones as a novel antibacterial chemotype with a polypharmacological mode of action, in which FabH, FphC, and AdhE play a central role.

Biochemical and Cellular Characterization of the Function of Fluorophosphonate-Binding Hydrolase H (FphH) in Staphylococcus aureus Support a Role in Bacterial Stress Response

“…That FphH can be targeted during the S. aureus life cycle by small molecules has been demonstrated. ,, Here we show that one of these small molecules, an oxadiazolone is indeed a strong inhibitor of FphH that covalently and therefore permanently binds to the active site serine residue, the proposed reaction mechanism of oxidazolones with serine hydrolases also appears to be confirmed by this study. Based on our atomic characterization of FphH, future structural studies of FphH in complex with small molecules bound to the active site could aid in the development of compounds that specifically target FphH.…”

“…Next we tested the inhibitory potential of a recently reported putative inhibitor for FphHthe covalent oxadiazole based inhibitor “compound 3” (Figure E) . The binding potential of compound 3 toward FphH had previously only been shown for a fluorescent probe version of compound 3 in S. aureus lysate.…”

“…The IC 50 was determined by using the GraphPad prism software. Compound 3 was synthesized as previously reported . The potential inhibitory effect of fusidic acid was measured similarly as described for compound 3 using 5 different concentrations (0.1 to 100 μM) and one h of preincubation, with no inhibition observed.…”

Chemoproteomic Approaches for Unraveling Prokaryotic Biology

“…FbhE, along with FphB and FphH, have also been identified as the cellular target of oxadiazolones that inhibit the growth of MRSA. [101] Active Fph homologs are found across numerous Staphylococcus spp., including Staphylococcus epidermis, [102] a commensal skin bacterium that regulates skin barrier integrity. [103] Accordingly, the effects of SA FphB inhibitors on commensal S. epidermis strains were characterized.…”

“…These FphE‐selective probes facilitated single‐cell characterization of activity and demonstrated FphE activity in different SA strains depended on growth conditions. FbhE, along with FphB and FphH, have also been identified as the cellular target of oxadiazolones that inhibit the growth of MRSA [101] …”