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The Diels–Alder (DA) reaction, a classic cycloaddition reaction involving a diene and a dienophile to form a cyclohexene, is among the most versatile organic reactions. Theories have predicted thermodynamically unfavorable DA reactions on pristine graphene owing to its low chemical reactivity. We hypothesized that metals like Ni could enhance the reactivity of graphene towards DA reactions through charge transfer. The results indeed showed that metal substrates enhanced the reactivity of graphene in the DA reactions with a diene, 2,3-dimethoxy butadiene (DMBD), and a dienophile, maleic anhydride (MAH), with the activity enhancement in the order of Ni > Cu, and both are more reactive than graphene supported on silicon wafer. The rate constants were estimated to be two times higher for graphene supported on Ni than on silicon wafer. The computational results support the experimentally obtained rate trend of Ni > Cu, both predicted to be greater than unsupported graphene, which is explained by the enhanced graphene–substrate interaction reflected in charge transfer effects with the strongly interacting Ni. This study opens up a new avenue for enhancing the chemical reactivity of pristine graphene through substrate selection. [ABSTRACT FROM AUTHOR]

The opportunistic pathogen Pseudomonas aeruginosa develops increasing resistance toward even the most potent antibiotics. Like other bacteria, the pathogen produces a number of virulence factors including metallophores, which constitute an important group. Pseudomonads produce the iron-chelating metallophore (siderophore) pyochelin, which, in addition to its iron-scavenging ability, is an effector for the transcriptional regulator PchR in its Fe III -bound form (ferripyochelin). In the present study, docking studies predicted a major ferripyochelin binding site in PchR, which prompted the exploration of nor-pyochelin analogues to produce tight binding to PchR, and thereby upregulation of the pyochelin metabolism. In addition, we investigated the effects of using the analogues to bind the antimicrobial cations Ga III and In III . Selected analogues of nor-pyochelin were synthesized, and their Ga III - and In III -based complexes were assessed for antimicrobial activity. The results indicate that the Ga III complexes inhibit the pathogens under iron-limited conditions, while the In III -based systems are more effective in iron-rich media. Several of the Ga III complexes were shown to be highly effective against a multidrug-resistant P. aeruginosa clinical isolate, with minimum inhibitory concentrations (MICs) of ≤1 μg/mL. Similarly, two of the In III -based systems were particularly effective against the isolate, with an MIC of 8 μg/mL. These results show high promise in comparison with other, traditionally potent antibiotics, as the compounds generally indicated low cytotoxicity toward mammalian cells. Preliminary mechanistic investigations using pseudomonal transposon mutants suggested that the inhibitory effects of the In III -based systems could be due to acute iron deficiency as a result of In III -bound bacterioferritin.

Real-time tracking of intracellular carbohydrates remains challenging. While click chemistry allows bio-orthogonal tagging with fluorescent probes, the reaction permanently alters the target molecule and only allows a single snapshot. Here, we demonstrate click-free mid-infrared photothermal (MIP) imaging of azide-tagged carbohydrates in live cells. Leveraging the micromolar detection sensitivity for 6-azido-trehalose (TreAz) and the 300-nm spatial resolution of MIP imaging, the trehalose recycling pathway in single mycobacteria, from cytoplasmic uptake to membrane localization, is directly visualized. A peak shift of azide in MIP spectrum further uncovers interactions between TreAz and intracellular protein. MIP mapping of unreacted azide after click reaction reveals click chemistry heterogeneity within a bacterium. Broader applications of azido photothermal probes to visualize the initial steps of the Leloir pathway in yeasts and the newly synthesized glycans in mammalian cells are demonstrated.

Drug repurposing efforts led to the discovery of bactericidal activity in auranofin, a gold-containing drug used to treat rheumatoid arthritis. Auranofin kills Gram-positive bacteria by inhibiting thioredoxin reductase, an enzyme that scavenges reactive oxygen species (ROS). Despite the presence of thioredoxin reductase in Gram-negative bacteria, auranofin is not always active against them. It is not clear whether the lack of activity in several Gram-negative bacteria is due to the cell envelope barrier or the presence of other ROS protective enzymes such as glutathione reductase (GOR). We previously demonstrated that chemical analogs of auranofin (MS-40 and MS-40S), but not auranofin, are bactericidal against the Gram-negative Burkholderia cepacia complex. Here, we explore the targets of auranofin, MS-40, and MS-40S in Burkholderia cenocepacia and elucidate the mechanism of action of the auranofin analogs by a genome-wide, randomly barcoded transposon screen (BarSeq). Auranofin and its analogs inhibited the B. cenocepacia thioredoxin reductase and induced ROS but did not inhibit the bacterial GOR. Genome-wide, BarSeq analysis of cells exposed to MS-40 and MS-40S compared to the ROS inducers arsenic trioxide, diamide, hydrogen peroxide, and paraquat revealed common and unique mediators of drug susceptibility. Furthermore, deletions of gshA and gshB that encode enzymes in the glutathione biosynthetic pathway led to increased susceptibility to MS-40 and MS-40S. Overall, our data suggest that the auranofin analogs kill B. cenocepacia by inducing ROS through inhibition of thioredoxin reductase and that the glutathione system has a role in protecting B. cenocepacia against these ROS-inducing compounds.IMPORTANCEThe Burkholderia cepacia complex is a group of multidrug-resistant bacteria that can cause infections in the lungs of people with the autosomal recessive disease, cystic fibrosis. Specifically, the bacterium Burkholderia cenocepacia can cause severe infections, reducing lung function and leading to a devastating type of sepsis, cepacia syndrome. This bacterium currently does not have an accepted antibiotic treatment plan because of the wide range of antibiotic resistance. Here, we further the research on auranofin analogs as antimicrobials by finding the mechanism of action of these potent bactericidal compounds, using a powerful technique called BarSeq, to find the global response of the cell when exposed to an antimicrobial., Competing Interests: The authors declare no conflict of interest.

In this work, we show that the addition of thiourea (TU) initiated broad-spectrum antimicrobial activity of otherwise inactive D-maltose-capped gold nanoclusters (AuNC-Mal). For example, AuNC-Mal/TU was effective against multidrug-resistant Pseudomonas aeruginosa with a minimum inhibitory concentration (MIC) of 1 μg mL -1 (2.5 μM [Au]) while having 30-60 times lower in vitro cytotoxicity against mammalian cells. The reaction of AuNC-Mal and TU generated the antimicrobial species of [Au(TU) 2 ] + and smaller AuNCs. TU increased the accumulation of Au in bacteria and helped maintain the oxidation state as Au I (vs. Au III ). The modes of action included the inhibition of thioredoxin reductase, interference with the Cu I regulation and depletion of ATP. Moreover, the antimicrobial activity did not change in the presence of colistin or carbonyl cyanide 3-chlorophenylhydrazone, suggesting that AuNC-Mal/TU was indifferent to the outer membrane barrier and to bacterial efflux pumps., (© 2023 Wiley-VCH GmbH.)

Auranofin, a gold(I)-complex with tetraacetylated thioglucose (Ac 4 GlcSH) and triethylphosphine ligands, is an FDA-approved drug used as an anti-inflammatory aid in the treatment of rheumatoid arthritis. In repurposing auranofin for other diseases, it was found that the drug showed significant activity against Gram-positive but was inactive against Gram-negative bacteria. Herein, the design and synthesis of gold nanoclusters (AuNCs) based on the structural motif of auranofin are reported. Phosphine-capped AuNCs are synthesized and glycosylated, yielding auranofin analogues with mixed triphenylphosphine monosulfonate (TPPMS)/Ac 4 GlcSH ligand shells. These AuNCs are active against both Gram-negative and Gram-positive bacteria, including multidrug-resistant pathogens. Notably, an auranofin analogue, a mixed-ligand 1.6 nm AuNC 4b, is more active than auranofin against Pseudomonas aeruginosa, while exhibiting lower toxicity against human A549 cells. The enhanced antibacterial activity of these AuNCs is characterized by a greater uptake of Au by the bacteria compared to Au I complexes. Additional factors include increased oxidative stress, moderate inhibition of thioredoxin reductase (TrxR), and DNA damage. Most intriguingly, the uptake of AuNCs are not affected by the bacterial outer membrane (OM) barrier or by binding with the extracellular proteins. This contrasts with Au I complexes like auranofin that are susceptible to protein binding and hindered by the OM barrier., (© 2021 Wiley-VCH GmbH.)