Monitoring host-pathogen interactions using chemical proteomics.

With the rapid emergence and the dissemination of microbial resistance to conventional chemotherapy, the shortage of novel antimicrobial drugs has raised a global health threat. As molecular interactions between microbial pathogens and their mammalian hosts are crucial to establish virulence, pathogenicity, and infectivity, a detailed understanding of these interactions has the potential to reveal novel therapeutic targets and treatment strategies. Bidirectional molecular communication between microbes and eukaryotes is essential for both pathogenic and commensal organisms to colonise their host. In particular, several devastating pathogens exploit host signalling to adjust the expression of energetically costly virulent behaviours. Chemical proteomics has emerged as a powerful tool to interrogate the protein interaction partners of small molecules and has been successfully applied to advance host-pathogen communication studies. Here, we present recent significant progress made by this approach and provide a perspective for future studies.

A chemical probe unravels the reactive proteome of health-associated catechols.

Catechol-containing natural products are common constituents of foods, drinks, and drugs. Natural products carrying this motif are often associated with beneficial biological effects such as anticancer activity and neuroprotection. However, the molecular mode of action behind these properties is poorly understood. Here, we apply a mass spectrometry-based competitive chemical proteomics approach to elucidate the target scope of catechol-containing bioactive molecules from diverse foods and drugs. Inspired by the protein reactivity of catecholamine neurotransmitters, we designed and synthesised a broadly reactive minimalist catechol chemical probe based on dopamine. Initial labelling experiments in live human cells demonstrated broad protein binding by the probe, which was largely outcompeted by its parent compound dopamine. Next, we investigated the competition profile of a selection of biologically relevant catechol-containing substances. With this approach, we characterised the protein reactivity and the target scope of dopamine and ten biologically relevant catechols. Strikingly, proteins associated with the endoplasmic reticulum (ER) were among the main targets. ER stress assays in the presence of reactive catechols revealed an activation of the unfolded protein response (UPR). The UPR is highly relevant in oncology and cellular resilience, which may provide an explanation of the health-promoting effects attributed to many catechol-containing natural products.

Eukaryotic catecholamine hormones influence the chemotactic control of Vibrio campbellii by binding to the coupling protein CheW.

SignificanceHost-emitted stress hormones significantly influence the growth and behavior of various bacterial species; however, their cellular targets have so far remained elusive. Here, we used customized probes and quantitative proteomics to identify the target of epinephrine and the α-adrenoceptor agonist phenylephrine in live cells of the aquatic pathogen Vibrio campbellii. Consequently, we have discovered the coupling protein CheW, which is in the center of the chemotaxis signaling network, as a target of both molecules. We not only demonstrate direct ligand binding to CheW but also elucidate how this affects chemotactic control. These findings are pivotal for further research on hormone-specific effects on bacterial behavior.

Fluorescent palladium(II) and platinum(II) NHC/1,2,3-triazole complexes: antiproliferative activity and selectivity against cancer cells.

Fluorescent Pd(ii) and Pt(ii) complexes bearing 4-methylene-7-methoxycoumarin (MMC) and 2,6-diispropylphenyl (Dipp) substituted NHC/1,2,3-triazole hybrid ligands are described. Depending on the reaction conditions two different ligand coordination modes are observed, i.e., bidentate solely coordinating via NHCs or tetradentate coordinating via NHCs and 1,2,3-triazoles. All Dipp substituted complexes show antiproliferative activity against cervix (HeLa) and breast (MCF-7) human carcinoma cells. The activity significantly depends on the coordination mode, with the tetradentate motif being notably more effective (HeLa: IC50 = 3.9 µM to 4.7 µM; MCF-7: IC50 = 2.07 µM to 2.35 µM). Amongst the MMC series, only the Pd(ii) complex featuring the bidentate coordination mode is active against HeLa (IC50 = 6.1 µM). In contrast to its structurally related Dipp derivative (SI = 0.6), it shows a high selectivity for HeLa (SI > 16) compared to healthy skin cells (HaCaT). According to fluorescence microscopy, this compound is presumably located in late endosomes or lysosomes.

The Natural Product Elegaphenone Potentiates Antibiotic Effects against Pseudomonas aeruginosa.

Natural products represent a rich source of antibiotics that address versatile cellular targets. The deconvolution of their targets via chemical proteomics is often challenged by the introduction of large photocrosslinkers. Here we applied elegaphenone, a largely uncharacterized natural product antibiotic bearing a native benzophenone core scaffold, for affinity-based protein profiling (AfBPP) in Gram-positive and Gram-negative bacteria. This study utilizes the alkynylated natural product scaffold as a probe to uncover intriguing biological interactions with the transcriptional regulator AlgP. Furthermore, proteome profiling of a Pseudomonas aeruginosa AlgP transposon mutant provided unique insights into the mode of action. Elegaphenone enhanced the elimination of intracellular P. aeruginosa in macrophages exposed to sub-inhibitory concentrations of the fluoroquinolone antibiotic norfloxacin.

Spontaneous Symmetry Breaking in the Formation of Supramolecular Polymers: Implications for the Origin of Biological Homochirality.

Aqueous solutions of the achiral, monomeric, nucleobase mimics (2,4,6-triaminopyrimidine, TAP, and a cyanuric acid derivative, CyCo6) spontaneously assemble into macroscopic homochiral domains of supramolecular polymers. These assemblies exhibit a high degree of chiral amplification. Addition of a small quantity of one handedness of a chiral derivative of CyCo6 generates exclusively homochiral structures. This system exhibits the highest reported degree of chiral amplification for dynamic helical polymers or supramolecular helices. Significantly, homochiral polymers comprised of hexameric rosettes with structural features that resemble nucleic acids are formed from mixtures of cyanuric acid (Cy) and ribonucleotides (l-, d-pTARC) that arise spontaneously from the reaction of TAP with the sugars. These findings support the hypothesis that nucleic acid homochirality was a result of symmetry breaking at the supramolecular polymer level.