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With the idea of exploiting metal templated C–S bond forming reactions to achieve modification of cysteines in bacterial proteins, a cyclometalated Au(III) compound was explored in a competitive chemoproteomic approach in S. aureus cell extracts. More than 100 ligandable cysteines were identified, of which more than 50% were not engaged by organic α-chloroacetamides in a previous study, indicating that organometallic compounds expand the ligandable space in bacteria. A selected interaction was validated using an enzyme activity assay, and intact protein mass spectrometry showed cysteine arylation of an unprecedented target. The obtained results demonstrate that this family of organogold compounds has potential for therapeutic protein targeting via selective, covalent modification of cysteine residues in bacteria. [ABSTRACT FROM AUTHOR]

In addition to their well-known role as stress-associated catecholamine hormones in animals and humans, epinephrine (EPI) and norepinephrine (NE) act as interkingdom signals between eukaryotic hosts and bacteria. However, the molecular basis of their effects on bacteria is not well understood. In initial phenotypic studies utilizing Vibrio campbellii as a model organism, we characterized the bipartite mode of action of catecholamines, which consists of promotion of growth under iron limitation and enhanced colony expansion on soft agar. In order to identify the molecular targets of the hormones, we designed and synthesized tailored probes for chemical proteomic studies. As the catechol group in EPI and NE acts as an iron chelator and is prone to form a reactive quinone moiety, we devised a photoprobe based on the adrenergic agonist phenylephrine (PE), which solely influenced colony expansion. Using this probe, we identified CheW, located at the core of the chemotaxis signaling network, as a major target. In vitro studies confirmed that EPI, NE, PE, and labetalol, a clinically applied antagonist, bind to purified CheW with affinity constants in the submicromolar range. In line with these findings, exposure of V. campbellii to these adrenergic agonists affects the chemotactic control of the bacterium. This study highlights an effect of eukaryotic signaling molecules on bacterial motility. [ABSTRACT FROM AUTHOR]

The development of novel anti-infectives requires unprecedented strategies targeting pathways which are solely present in pathogens but absent in humans. Following this principle, we developed inhibitors of lipoic acid (LA) salvage, a crucial pathway for the survival of LA auxotrophic bacteria and parasites but non-essential in human cells. An LA-based probe was selectively transferred onto substrate proteins via lipoate protein ligase (LPL) in intact cells, and their binding sites were determined by mass spectrometry. Probe labeling served as a proxy of LPL activity, enabling in situ screenings for cell-permeable LPL inhibitors. Profiling a focused compound library revealed two substrate analogs (LAMe and C3) as inhibitors, which were further validated by binding studies and co-crystallography. Importantly, LAMe exhibited low toxicity in human cells and achieved killing of Plasmodium falciparum in erythrocytes with an EC 50 value of 15 μM, making it the most effective LPL inhibitor reported to date., (© 2023 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)