Cystobactamids are nonribosomal peptide natural products that function as DNA gyrase inhibitors, exhibiting significant antibacterial activity. They are isolated from Cystobacter sp. Cbv34 and contain various alkoxy groups on para-aminobenzoic acid moieties, which are believed to play a crucial role in antibacterial functions. The alkoxy groups are generated by iterative methylations on a methoxy group by the cobalamin (Cbl)-dependent radical S-adenosylmethionine (SAM) enzyme CysS. CysS catalyzes up to three methylations to give ethoxy, isopropoxy, sec-butoxy, and tert-butoxy groups. For each methylation, CysS uses a ping-pong mechanism in which two molecules of SAM are consumed. One SAM is used to methylate cob(I)alamin, while another generates a 5'-deoxyadenosyl 5'-radical to initiate substrate methylation. However, little is known about how the enzyme promotes both Cbl methylation and iterative substrate methylation, which occur by polar SN2 and radical processes, respectively. Here, we report three X-ray crystal structures of a homolog of CysS from Corallococcus sp. CA054B. Two were determined in the presence of methoxy- and ethoxy-containing substrates, showing how CysS accommodates substrates and products during iterative methylation. The third structure, determined in the absence of a substrate, exhibits structural changes that reorient the SAM's conformation to allow for the methylation of cob(I)alamin.

The Calophyllum species have gained a lot of attention for their structurally diverse secondary metabolites with potential biological activities, including antibacterial scaffolds. A detailed study done on the phytochemical profile of the Calophyllum nodosum stem bark has led to the isolation of three xanthones, trapezifolixanthone (1), caloxanthone C (2), 1-hydroxy-7-methoxyxanthone (3), and a cyclic ester, canumolactone (4). Their structures were characterised using spectroscopic techniques like NMR, MS, and IR, and the spectra were confirmed with the previous literature. The pharmacokinetic properties of the compounds were predicted using SwissADME and cross-validated using pkCSM, while molecular docking simulation against bacterial DNA gyrase was performed using Autodock Vina. All isolated compounds met the drug-likeness requirements under Lipinski’s rule, according to the ADMET predictions, showing favourable oral and gastrointestinal bioavailability and absorption. Canumolactone (4), in particular, showed the best combination of solubility, clearance, and the least amount of CYP liabilities. The molecular docking simulation revealed that trapezifolixanthone (1) gave the strongest binding affinity to DNA gyrase among all isolated compounds, surpassing the binding affinity of the standard inhibitor, BDBM50198240. The current study presents the first combined ADMET predictions and molecular docking analysis for compounds isolated from C. nodosum. The findings provide preliminary evidence of their potential as drug-like scaffolds for future pharmacological development.

Machine learning and generative AI in the rational design of DNA gyrase-targeted antibacterials.

In silico identification of potential inhibitors of Mycobacterium tuberculosis DNA gyrase from Phytoconstituents of Indian medicinal plants.

Molecular modeling and optimization of pyranoquinolinone derivatives as potential DNA gyrase inhibitors for typhoid fever treatment: A Comprehensive QSAR, molecular docking, ADMET, and molecular dynamics simulation studies

Cystobactamids are non-ribosomal peptide natural products that function as DNA gyrase inhibitors, exhibiting significant antibacterial activity. They are isolated from Cystobacter sp. Cbv34 and contain various alkoxy groups on para-aminobenzoic acid moieties, which are believed to play a crucial role in antibacterial functions. The alkoxy groups are generated by iterative methylations on a methoxy group by the cobalamin (Cbl)-dependent radical S-adenosylmethionine (SAM) enzyme CysS. CysS catalyzes up to three methylations to give ethoxy, isopropoxy, sec-butoxy, and tert-butoxy groups. For each methylation, CysS uses a ping-pong mechanism in which two molecules of SAM are consumed. One SAM is used to methylate cob(I)alamin, while another generates a 5′-deoxyadenosyl 5′-radical to initiate substrate methylation. However, little is known about how the enzyme promotes both Cbl methylation and iterative substrate methylation, which occur by polar SN2 and radical processes, respectively. Here, we report three X-ray crystal structures of a homolog of CysS from Corallococcus sp. CA054B. Two were determined in the presence of methoxy- and ethoxy-containing substrates, showing how CysS accommodates substrates and products during iterative methylation. The third structure, determined in the absence of a substrate, exhibits structural changes that reorient the SAM’s conformation to allow for the methylation of cob(I)alamin.

The emergence of drug resistance remains a major challenge in the treatment of tuberculosis and other mycobacterial infections. To combat the rise in resistance, strategies that reduce the frequency of resistance mutations are urgently needed. Verapamil is a small-molecule compound that can enhance the potency of companion drugs in combination regimen. Here, we investigate if verapamil can decrease the resistance frequency of antimycobacterial drugs. The results show that verapamil significantly reduces the resistance frequency of multiple antimycobacterial agents, including the DNA gyrase inhibitor moxifloxacin, the protein synthesis inhibitor streptomycin, and the RNA polymerase inhibitor rifampicin in Mycobacterium smegmatis. The presence of point mutations in the target was confirmed for moxifloxacin-resistant M. smegmatis. Suppression of resistance evolution against moxifloxacin by verapamil was also found in the slow-growing, pathogenic mycobacteria M. avium and M. tuberculosis. Real-time qPCR analysis in M. smegmatis showed that verapamil treatment downregulates the expression of multiple efflux pump genes and upregulates DNA repair genes. These findings suggest that verapamil exerts a dual role by interfering with efflux pump functionality and by reducing the probability of chromosomal mutations. The combination of these properties may underlie the promise of verapamil as adjuvant to enhance the effectiveness of current antimycobacterial chemotherapy.

In-silico design of some tetrazoloquinoline analogs as potent inhibitors of DNA gyrase of Salmonella typhi: QSAR modeling, molecular docking, MD simulation, and ADMET profiling

Endophytic Aspergillus sp. from Phragmites australis: A source of DNA gyrase inhibitors with antibiofilm, antioxidant, and cytotoxic potentials: in vitro supported by in silico

New quinazoline analogues as Mtb DNA gyrase inhibitors: Design, synthesis, antimycobacterial evaluation and computational studies

Covering: 2014/2015 up to 2025.The global rise of antimicrobial resistance imposes a strong demand to develop new antibacterial drugs, and microbes have been a prime source for their discovery. Albicidins and cystobactamids, isolated from xanthomonadaceae and myxococcaceae, respectively, span a novel class of oligoarylamide antibiotics with a unique chemical scaffold featured by para-aminobenzoic acid building blocks. Both compounds exhibit broad spectrum and potent activity against Gram-positive and Gram-negative pathogens through inhibiting DNA gyrase and topoisomerase IV. This article summarizes the insights gained on this class since its initial disclosure in 2014/2015 up to 2025. It discusses natural derivatives, their biosynthesis and chemical synthesis, the unique binding mode to DNA gyrase, and systematic medicinal chemistry programs with >700 analogs that led to resistance-breaking antibiotics with in vivo efficacy. The review illustrates the importance of natural product research to address the global need for new antibiotics.

To address the increasing Antimicrobial Resistance (AMR), we developed a library of triazole-tethered tetrazole derivatives using a multicomponent synthetic click chemistry strategy. It is well known that combining two or more types of pharmacophores into one molecule could afford a new entity with varied bioactivities. Considering this, the final products (6a–6o) were synthesized in excellent yields and were duly characterized using spectrometric analysis, including NMR and HRMS. To rationalize their biological attributes, synthetics were tested using different pathogenic microbial strains (S. aureus (ATCC 25923), S. epidermidis (ATCC 35984), E. coli (ATCC 25922), A. hydrophila (ATCC 7966), P. aeruginosa (ATCC 27853), S. typhi (Clinical isolate), S. typhimurium (Clinical isolate)). The antimicrobial potential (MIC µg/mL) of compounds compared to positive control ciprofloxacin revealed that compounds 6a, 6b, 6c, 6d, 6e, 6g, 6h, 6j, 6l, and 6m exhibited significant antibacterial activity with MIC 1.56-3.12 µg/mL in vitro compared to the ciprofloxacin against Gram-positive and Gram-negative bacterial strains. The molecules were further corroborated rationally using molecular modelling and dynamics analysis to assess their binding affinity with DNA gyrase. The study established that 6g and 6e possess a high affinity within the gyrase, as revealed by molecular docking analysis compared to ciprofloxacin. The molecular dynamics analysis for 6g revealed a stable conformation within the protein domain during the simulation period. The present work thus opens up the possibility of further exploring the utility of 6g and 6e in delineating their DNA gyrase binding biologically and deducing their mechanistic interventions. The work may further be expanded to recruit more pathogenic-resistant strains, and the inhibitory potential of the compounds may further be analysed.Supplementary InformationThe online version contains supplementary material available at 10.1038/s41598-025-15919-4.

Expanding antimicrobial chemotypes: indole-based DNA gyrase inhibitors with potential dual mechanism against multidrug-resistant bacteria.

Synthesis and Molecular Docking Study of 2-Aroyl-3-phosphonylbenzofurans as Bacterial DNA Gyrase Inhibitors

Antimicrobial resistance poses a critical challenge to global public health, exacerbating morbidity and mortality associated with bacterial infections. This study addresses the urgent need for novel antibacterial agents by exploring the design and synthesis of quinazoline-piperazine phosphorodiamidate hybrids (6a-g) as potential DNA gyrase inhibitors. Antibacterial activity was evaluated using the agar well diffusion method, revealing significant inhibition zones for compounds 6f, 6g, 6a, and 6c compared to the standard drug Amoxyclav. Minimum inhibitory concentration (MIC) measurements further supported the potent antibacterial effects of these compounds. Additionally, compounds 6f, 6g, and 6a exhibited notable antifungal activity superior to Fluconazole. Molecular docking simulations against DNA gyrase demonstrated strong binding affinities of compounds 6f and 6a with dock scores surpassing that of a standard antibiotic, ciprofloxacin. Detailed analysis of binding interactions highlighted key residues involved in stabilizing the ligand-protein complexes, providing insights into their mechanism of action. Furthermore, in silico ADMET prediction studies revealed that the targeted analogs satisfied the drug like characteristics of CNS acting drugs against antimicrobial diseases.

The increasing use of antibiotics coupled with the lack of innovative and effective antimicrobial agents has increased the development of antimicrobial resistance (AMR) worldwide. To overcome the AMR-associated prolonged disease duration and increased mortality rates, new antimicrobial agents are in high demand. In this context, hydrazone and oxadiazole derivatives are endowed with remarkable biocidal activity, becoming profitable scaffolds in the design of antimicrobial candidates. In this study, the antimicrobial effects of N-acyl hydrazones 1-15 and 2,5-disubstituted 1,3,4- oxadiazoles 16-27 against Escherichia coli ATCC 25922, Staphylococcus aureus ATCC 29213, Bacillus subtilis ATCC 6633, and clinically isolated Shigella sonnei, Klebsiella pneumoniae, and Candida albicans were evaluated. For this purpose, Kirby-Bauer disc diffusion and MIC tests were carried out, indicating that most of these compounds were active against tested microorganisms. Particularly, several compounds proved active against E. coli, whereas S. aureus showed higher resistance. The genotoxic potential of most active compounds was determined by in vitro alkaline comet assay, and they were found to be non-toxic at studied concentrations. Finally, molecular docking and dynamics (MD) studies identified four compounds as potential inhibitors of bacterial DNA gyrase B (GyrB). Further exploration of molecular determinants revealed favourable drug-like properties, highlighting the potential of these molecules for subsequent hit-to-lead optimization studies.

The emergent antimicrobial resistance (AMR) has required generating new antibacterial substances with distinct functions. DNA gyrase- particularly the subunit GyrB has been identified as a favorable bacterial target because it contains a highly conserved ATP binding site, and is required in supercoiling of DNA. Broad-spectrum pharmacological agents flaunted by thiazole derivatives have been proven to be highly antibacterial agents when rationally prepared to target gyrB. The review indicates the use of modern computational methods of drug design--QSAR modeling, molecular docking, molecular dynamics and ADMET prediction--to identify and optimize thiazole-based DNA gyrase inhibitors. Such in silico approaches enabled weeks worth of candidate screening in analogy of thiazole compounds with high binding affinities, good pharmacokinetics and low toxicity. Remarkably some of the lead substances showed good inhibition of bacteria in an in vivo setting with low bacterial load and gentle side effects on animal subjects. Not only that, the thiazole derivatives also demonstrated the capability of overcoming the existing resistance mechanisms like GyrA mutations and efflux pump by inhibiting a region less prone to mutations in GyrB. The combination of computer and experimental-based methods is not only advancing the process of drug discovery but also helping in enabling the design of resistance evading antibacterial agents with structural novelty. Therefore, thiazole-based inhibitors are an interesting opportunity in next-generation antibiotics with the constantly growing AMR problems around the globe.

Design and synthesis of β-carboline-benzofuran based hybrids as antibacterial agents against Staphylococcus aureus.

Abstract A novel series of 3‐methylquinazolin‐4‐ones 3, 5, 7, 8, and 10 linked to aryl hydrazone, pyrazole, pyridazine, and thiazole through thio linker was generated and described in the present work, starting from quinazoline acetonitrile 1. Spectroscopic and microanalytical data were used to investigate the newly generated materials. The antimicrobial efficacy of these molecules was tested using amoxicillin and ketoconazole, which are standard antimicrobial agents. The new compounds were examined against several bacterial and fungal microorganisms. Pyrazole analogue 5b showed excellent potency rather than 5a, resembling that of the reference amoxicillin (MIC = 1.95, 3.9, 0.98, and 0.49 µM against S. aureus, E. faecalis, K. pneumoniae, and P. aeruginosa, respectively). Also, derivative 5b gave moderate activity against fungi A. niger and C. albicans (MIC = 15.63 and 125 µM, respectively) comparing with the standard ketoconazole (MIC = 7.81 and 62.5 µM, respectively). The above derivatives were evaluated for inhibition of DNA gyrase and topoisomerase IV of S. aureus in relation to the common drug ciprofloxacin. Although the tested derivatives 5a, b had a potential suppressive effect against S. aureus DNA gyrase when compared to the standard used (IC50 = 8.15 ± 0.55, 6.44 ± 0.30 and 5.72 ± 0.13 µM, respectively), they revealed a significant decrease in the suppressive effect on topoisomerase IV compared to standard drugs, with (IC50 range 61.58–95.40 µM, IC50 ciprofloxacin = 14.88 ± 0.25 µM). Utilizing molecular docking technology, the more potent molecules, 5a and 5b, were further examined for the ability to attach to the required DNA gyrase of S. aureus. Hydrogen bonding interactions between their structures and protein residues such as Pro1080 and His1081 were observed in the DNA gyrase active pocket of S. aureus. Both pyrazoles 5a and 5b appear to have good prospects for further research progress and improvement, according to computational evaluation of physicochemical parameters and ADMET.

DNA gyrase is a type II topoisomerase enzyme that can cause negative supercoiling in DNA by using the energy produced by ATP hydrolysis. There are two main types of topoisomerases: type I and type II. Type I enzymes cut a single strand of DNA and are further classified as type IA if they connect to a 5′ phosphate of DNA, or type IB if they link to a 3′ phosphate. Type II topoisomerases break both strands, creating a staggered double-strand break. Antimicrobial resistance is a major concern for the global healthcare system. Resistance is the ability of microorganisms to neutralize and withstand antimicrobial drugs previously used to treat microbial infections. Some known classes of DNA gyrase inhibitors are coumarins, cyclothialidines, and quinolones. Antimicrobial medicines such as quinolones have been widely used to treat microbiological diseases. However, the increased use of quinolones has led to the emergence of quinolone-resistant bacteria, which poses a serious risk to public health. Microorganisms can cause resistance due to changes in the target enzymes, DNA gyrase, and topoisomerase IV, which are responsible for transcription and DNA replication. Additionally, differences in drug entry and efflux may also play a role in resistance. Plasmids that produce the Qnr protein can mediate resistance to quinolones by protecting the quinolone targets from inhibition. This review aims to revolutionize the discovery of quinolone-based antibiotics, specifically targeting DNA gyrase, a critical enzyme in bacterial DNA replication, to enhance the efficacy and specificity of anti-microbail agents against microbial infections.