Enzymes belonging to the shikimate pathway have long been considered promising targets for antibacterial drugs because they have no counterpart in mammals and are essential for bacterial growth and ...virulence. However, despite decades of research, there are currently no clinically relevant antibacterial drugs targeting any of these enzymes, and there are legitimate concerns about whether they are sufficiently druggable, i.e., whether they can be adequately modulated by small and potent drug-like molecules. In the present work, in silico analyses combining evolutionary conservation and druggability are performed to determine whether these enzymes are candidates for broad-spectrum antibacterial therapy. The results presented here indicate that the substrate-binding sites of most enzymes in this pathway are suitable drug targets because of their reasonable conservation and druggability scores. An exception was the substrate-binding site of 3-deoxy-
-arabino-heptulosonate-7-phosphate synthase, which was found to be undruggable because of its high content of charged residues and extremely high overall polarity. Although the presented study was designed from the perspective of broad-spectrum antibacterial drug development, this workflow can be readily applied to any antimicrobial target analysis, whether narrow- or broad-spectrum. Moreover, this research also contributes to a deeper understanding of these enzymes and provides valuable insights into their properties.
Antimicrobial resistance is a global health threat that requires innovative strategies against drug-resistant bacteria. Our study focuses on enoyl-acyl carrier protein reductases (ENRs), in ...particular FabI, FabK, FabV, and InhA, as potential antimicrobial agents. Despite their promising potential, the lack of clinical approvals for inhibitors such as triclosan and isoniazid underscores the challenges in achieving preclinical success. In our study, we curated and analyzed a dataset of 1412 small molecules recognized as ENR inhibitors, investigating different structural variants. Using advanced cheminformatic tools, we mapped the physicochemical landscape and identified specific structural features as key determinants of bioactivity. Furthermore, we investigated whether the compounds conform to Lipinski rules, PAINS, and Brenk filters, which are crucial for the advancement of compounds in development pipelines. Furthermore, we investigated structural diversity using four different representations: Chemotype diversity, molecular similarity, t-SNE visualization, molecular complexity, and cluster analysis. By using advanced bioinformatics tools such as matched molecular pairs (MMP) analysis, machine learning, and SHAP analysis, we were able to improve our understanding of the activity cliques and the precise effects of the functional groups. In summary, this chemoinformatic investigation has unraveled the FAB inhibitors and provided insights into rational antimicrobial design, seamlessly integrating computation into the discovery of new antimicrobial agents.
Optimized conditions for the synthesis of fully deprotected (2R)-aziridine containing dipeptides are described. Preparation of fully protected N- and C- terminal aziridine containing dipeptides was ...found to be straightforward and high yielding for the majority of compounds, whereas their full deprotection was possible only for C-terminal analogs. Deprotection of N-terminal derivatives using standard procedures of peptide chemistry was found difficult providing only mixtures of unidentifiable products. The described molecules have potential as building blocks in synthetic chemistry, in the chemical biology arena, as covalent modifiers, and as biomarkers.
Despite the great importance of β-lactam antibiotics, there is still a limited number of synthetic approaches for the formation of β-lactam⁻containing dipeptides. In this study, we report upon the ...stereoselective preparation of β-lactam⁻containing pseudopeptides, where different reaction conditions and NH₂ protective groups were tested to obtain compounds that contain 3-amino-azetidin-2-one. We demonstrate that the protective group is essential for the outcome of the reaction. Successful implementation of dibenzyl-protected serine-containing dipeptides through the Mitsunobu reaction can provide the desired products at high yields and stereoselectivity.
Bacterial resistance is an increasing threat to healthcare systems, highlighting the need for discovering new antibacterial agents. An established technique, fragment-based drug discovery, was used ...to target a bacterial enzyme Ddl involved in the biosynthesis of peptidoglycan. We assembled general and focused fragment libraries that were screened in a biochemical inhibition assay. Screening revealed a new fragment-hit inhibitor of DdlB with a Ki value of 20.7 ± 4.5 µM. Binding to the enzyme was confirmed by an orthogonal biophysical method, surface plasmon resonance, making the hit a promising starting point for fragment development.
MurA catalyzes the first step of peptidoglycan (PG) biosynthesis and is a validated target for the development of new antimicrobial agents. In this study, a library of 49 plant flavonoids and their ...synthetic derivatives were evaluated for their inhibitory properties against MurA fromEscherichia coli. The compounds were tested with and without preincubation and with the addition of DTT to understand the mechanism of inhibition. Thirteen compounds were identified as reversible, time-dependent inhibitors, with inhibition levels ranging from 480 nM to 57 μM, and ampelopsin as the most potent compound. To investigate the major pharmacophore elements responsible for the activity, 2D-QSAR and docking analyzes were performed. The results showed that the catechol moiety and an additional aromatic system were the most important features contributing to the activity of the compounds. However, most of the compounds did not show antibacterial activity againstE. coli andStaphylococcus aureusstrains, suggesting that their inhibitory activity against MurA may not be strong enough to induce antibacterial effects. Nevertheless, our results suggest that flavonoids are a promising starting point to develop new inhibitors of MurA and show great potential for further steps in drug development.
The use of green extraction methods that meet the criteria of sustainable and environmentally friendly technologies has been increasing in recent decades due to their many benefits. In this respect, ...extracts obtained using subcritical water are also gaining increased attention because of their potential antioxidant and antimicrobial properties. Their antimicrobial activity is mainly due to the presence of various polyphenolic compounds. Although the exact mechanism of the antibacterial action of polyphenolic compounds has not yet been fully investigated and described, polyphenols are known to affect the bacterial cell at several cellular levels; among other things, they cause changes and ruptures in the cell membranes of the bacterial cell, affect the inactivation of bacterial enzymes and damage bacterial DNA. The difference in the strength of the antimicrobial activity of the extracts is most likely a result of differences in their lipophilicity and in the number and position of hydroxyl groups and double bonds in the chemical structure of polyphenols. By changing the extraction conditions, especially the temperature, during subcritical water extraction, we affect the solubility of the compounds we want to extract. In general, as the temperature increases, the solubility of polyphenolic compounds also increases, and the reduction of the surface tension of subcritical water at higher temperatures also enables faster dissolution of polyphenolic compounds. Different bacterial strains have different sensitivity to different extracts. However, extracts obtained with subcritical water extraction demonstrate strong antimicrobial activity compared to extracts obtained with conventional methods.
Quorum sensing (QS), a bacterial communication strategy, has been recognized as one of the control mechanisms of virulence in bacteria. Thus, targeting QS offers an interesting opportunity to impair ...bacterial pathogenicity and develop antivirulence agents. Aiming to enhance the discovery of QS inhibitors, we developed a bioreporter Escherichia coli JW5505 pET‐Plsrlux and set up a cell‐based assay for identifying inhibitors of autoinducer‐2 (AI‐2)‐mediated QS. A comparative study on the performance of target‐ versus cell‐based assays was performed, and 91 compounds selected with the potential to target the ATP binding pocket of LsrK, a key enzyme in AI‐2 processing, were tested in an LsrK inhibition assay, providing 36 hits. The same set of compounds was tested by the AI‐2‐mediated QS interference assay, resulting in 24 active compounds. Among those, six were also found to be active against LsrK, whereas 18 might target other components of the pathway. Thus, this AI‐2‐mediated QS interference cell‐based assay is an effective tool for complementing target‐based assays, yet also stands as an independent assay for primary screening.
Exploring quorum sensing inhibition: Target‐ and bioreporter‐based approaches were used in parallel to screen 91 compounds targeted at the ATP binding pocket of LsrK, a kinase that functions in the autoinducer‐2 pathway. Six compounds active against LsrK also inhibited QS activation in the novel E. coli JW5503 pET‐Plsrlux bioreporter. The cell‐based setup provided 18 additional hit compounds with unknown target.
Various organic impurities (starting materials, reagents, intermediates, degradation products, by-products, and side products) could be present in active pharmaceutical ingredients affecting their ...qualities, safeties, and efficacies. Herein, we present the efficient syntheses of two United States Pharmacopeia impurities of an antidiabetic drug sitagliptin, a potent and orally active dipeptidyl peptidase IV inhibitor: 3-desamino-2,3-dehydrositagliptin and 3-desamino-3,4-dehydrositagliptin. Our three-step synthetic approach is based on the efficient cobalt-catalyzed cross-coupling reaction of 1-bromo-2,4,5-trifluorobenzene and methyl 4-bromocrotonate in the first step, followed by hydrolysis of corresponding ester with 3 M HCl to (E)-(2,4,5-trifluorophenyl)but-2-enoic acid in high overall yield, whereas the reaction with 3 M NaOH resulted in the carbon–carbon double bond regio-isomerization and hydrolysis to give the (E)-(2,4,5-trifluorophenyl)but-3-enoic acid in 92% yield. Both acid derivatives were converted to title compounds via the amide bond formation with 3-(trifluoromethyl)-5,6,7,8-tetrahydro-1,2,4triazolo4,3-apyrazine. Extensive screening of coupling/activation reagents, bases, and solvents reviled that the amide bond is formed the most efficiently using the (COCl)2/Et3N in THF or alternatively EDC/NMM/(DMAP or HOBt) in DMF obtaining the title compounds in 68–76% yields and providing the overall yields for the three-step process in the range of 57–64% on a gram scale. The presented study also demonstrates the importance of a proper selection of solvent, base, and coupling/activating reagent for amide bond formation using Michael acceptor-type allylbenzene derivatives as coupling partners to minimize the carbon–carbon double bond regio-isomerization.
Enzymes MurA and MurF, involved in bacterial cell wall synthesis, have been validated as targets for the discovery of novel antibiotics. A panel of plant-origin antibacterial diterpenes and synthetic ...analogs derived therefrom were investigated for their inhibitory properties on these enzymes from
and
. Six compounds were proven to be effective for inhibiting MurA from both bacteria, with IC
values ranging from 1.1 to 25.1 µM. To further mechanistically investigate the nature of binding and to explain the activity, these compounds were docked into the active site of MurA from
. The aromatic ring of the active compounds showed a T-shaped π-π interaction with the phenyl ring of Phe328, and at least one hydrogen bond was formed between the hydroxy groups and Arg120 and/or Arg91. The results disclosed here establish new chemical scaffolds for the development of novel entities targeting MurA as potential antibiotics to combat the threat of pathogenic bacteria, particularly resistant strains.
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