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Nucleic Acids Research Feb 2020Negative supercoiling by DNA gyrase is essential for maintaining chromosomal compaction, transcriptional programming, and genetic integrity in bacteria. Questions remain...
Negative supercoiling by DNA gyrase is essential for maintaining chromosomal compaction, transcriptional programming, and genetic integrity in bacteria. Questions remain as to how gyrases from different species have evolved profound differences in their kinetics, efficiency, and extent of negative supercoiling. To explore this issue, we analyzed homology-directed mutations in the C-terminal, DNA-wrapping domain of the GyrA subunit of Escherichia coli gyrase (the 'CTD'). The addition or removal of select, conserved basic residues markedly impacts both nucleotide-dependent DNA wrapping and supercoiling by the enzyme. Weakening CTD-DNA interactions slows supercoiling, impairs DNA-dependent ATP hydrolysis, and limits the extent of DNA supercoiling, while simultaneously enhancing decatenation and supercoil relaxation. Conversely, strengthening DNA wrapping does not result in a more extensively supercoiled DNA product, but partially uncouples ATP turnover from strand passage, manifesting in futile cycling. Our findings indicate that the catalytic cycle of E. coli gyrase operates at high thermodynamic efficiency, and that the stability of DNA wrapping by the CTD provides one limit to DNA supercoil introduction, beyond which strand passage competes with ATP-dependent supercoil relaxation. These results highlight a means by which gyrase can evolve distinct homeostatic supercoiling setpoints in a species-specific manner.
Topics: Adenosine Triphosphate; Catalysis; Chromosomes, Bacterial; DNA Gyrase; DNA, Bacterial; DNA, Superhelical; Escherichia coli; Models, Molecular; Mutation; Nucleic Acid Conformation; Protein Binding; Protein Domains
PubMed: 31950157
DOI: 10.1093/nar/gkz1230 -
Bioorganic Chemistry May 2023New 6,7-dimethylquinoxalin-2(1H)-one and hydrazineylidene thiazol-4-one derivatives were synthesized, and evaluated for their in vitro antimicrobial activity. The...
New 6,7-dimethylquinoxalin-2(1H)-one and hydrazineylidene thiazol-4-one derivatives were synthesized, and evaluated for their in vitro antimicrobial activity. The obtained results revealed marked antimicrobial potential against four bacterial, and two fungal strains. Both 6,7-dimethyl-3-(2-(4-nitrophenyl)-2-oxoethyl)quinoxalin-2(1H)-one (4d), and 2-(2-(9H-fluoren-9-ylidene)hydrazineyl)-5-(2-(p-tolyl)hydrazineylidene)thiazol-4(5H)-one (11b) displayed significant antibacterial and antifungal activities having MIC ranges (1.98-15.6 mg/mL) and (1.98-3.9 mg/mL) compared to Tetracycline and Amphotericin B as standard drugs. In addition, they showed noticeable inhibitory activity against DNA gyrase enzyme. Interestingly the thiazole derivative (11b) showed marked inhibitory activity against DNA gyrase with IC = 7.82 ± 0.45 μM better than that of ciprofloxacin. The time-kill kinetics profile of the most active compounds against S. aureus and E. coli microorganisms displayed both concentration dependent and time dependent reduction in the number of viable cells. Furthermore, molecular docking study of both compounds in the DNA gyrase binding site was performed, showing agreement with the in vitro inhibitory activities.
Topics: DNA Gyrase; Thiazoles; Molecular Docking Simulation; Staphylococcus aureus; Escherichia coli; Anti-Bacterial Agents; Microbial Sensitivity Tests; Topoisomerase II Inhibitors; Molecular Structure
PubMed: 36842318
DOI: 10.1016/j.bioorg.2023.106433 -
Proceedings of the National Academy of... Mar 2021DNA gyrase, a type II topoisomerase, introduces negative supercoils into DNA using ATP hydrolysis. The highly effective gyrase-targeted drugs, fluoroquinolones (FQs),...
DNA gyrase, a type II topoisomerase, introduces negative supercoils into DNA using ATP hydrolysis. The highly effective gyrase-targeted drugs, fluoroquinolones (FQs), interrupt gyrase by stabilizing a DNA-cleavage complex, a transient intermediate in the supercoiling cycle, leading to double-stranded DNA breaks. MfpA, a pentapeptide-repeat protein in mycobacteria, protects gyrase from FQs, but its molecular mechanism remains unknown. Here, we show that MfpA (MsMfpA) inhibits negative supercoiling by gyrase (Msgyrase) in the absence of FQs, while in their presence, MsMfpA decreases FQ-induced DNA cleavage, protecting the enzyme from these drugs. MsMfpA stimulates the ATPase activity of Msgyrase by directly interacting with the ATPase domain (MsGyrB47), which was confirmed through X-ray crystallography of the MsMfpA-MsGyrB47 complex, and mutational analysis, demonstrating that MsMfpA mimics a T (transported) DNA segment. These data reveal the molecular mechanism whereby MfpA modulates the activity of gyrase and may provide a general molecular basis for the action of other pentapeptide-repeat proteins.
Topics: Adenosine Triphosphatases; Bacterial Proteins; Crystallography, X-Ray; DNA Cleavage; DNA Gyrase; Molecular Mimicry; Monomeric GTP-Binding Proteins; Mycobacterium; Protein Conformation
PubMed: 33836580
DOI: 10.1073/pnas.2016705118 -
Archiv Der Pharmazie Jan 2022A series of 3-[(1H-pyrazol-3-yl)imino]indolin-2-one derivatives were designed using the molecular hybridization method, characterized using different spectroscopic... (Comparative Study)
Comparative Study
A series of 3-[(1H-pyrazol-3-yl)imino]indolin-2-one derivatives were designed using the molecular hybridization method, characterized using different spectroscopic techniques, and evaluated for their in vitro antimicrobial activity. Most of the target compounds demonstrated good to moderate antimicrobial activity compared with ciprofloxacin and fluconazole. Four compounds (8b, 9a, 9c, and 10a) showed encouraging results, with minimal inhibitory concentration (MIC) values (53.45-258.32 µM) comparable to those of norfloxacin (100.31-200.63 µM) and ciprofloxacin (48.33-96.68 µM). Noticeably, the four derivatives revealed excellent bactericidal and fungicidal activities, except for the bacteriostatic potential of compounds 8b and 9a against Escherichia coli and Staphylococcus aureus, respectively. The time-killing kinetic study against S. aureus confirmed the efficacy of these derivatives. Furthermore, two of the four promising derivatives, 9a and 10a, could prevent the formation of biofilms of S. aureus without affecting the bacterial growth at low concentrations. A combination study with seven commercial antibiotics against the multidrug-resistant bacterium P. aeruginosa showed a notable reduction in the antibiotic MIC values, represented mainly through a synergistic or additive effect. The enzymatic assay implied that the most active derivatives had inhibition potency against DNA gyrase comparable to that of ciprofloxacin. Molecular docking and density functional theory calculations were performed to explore the binding mode and study the reactivity of the promising compounds.
Topics: Anti-Infective Agents; Ciprofloxacin; DNA Gyrase; Indoles; Microbial Sensitivity Tests; Models, Molecular; Molecular Docking Simulation; Norfloxacin; Pyrazoles; Structure-Activity Relationship; Topoisomerase II Inhibitors
PubMed: 34747519
DOI: 10.1002/ardp.202100266 -
Antimicrobial Agents and Chemotherapy Sep 2022Tricyclic pyrrolopyrimidines (TPPs) are a new class of antibacterials inhibiting the ATPase of DNA gyrase. TPP8, a representative of this class, is active against...
Tricyclic pyrrolopyrimidines (TPPs) are a new class of antibacterials inhibiting the ATPase of DNA gyrase. TPP8, a representative of this class, is active against Mycobacterium abscessus . Spontaneous TPP8 resistance mutations mapped to the ATPase domain of M. abscessus DNA gyrase, and the compound inhibited DNA supercoiling activity of recombinant M. abscessus enzyme. Further profiling of TPP8 in macrophage and mouse infection studies demonstrated proof-of-concept activity against M. abscessus and .
Topics: Adenosine Triphosphatases; Animals; Anti-Bacterial Agents; DNA Gyrase; Mice; Microbial Sensitivity Tests; Mycobacterium Infections, Nontuberculous; Mycobacterium abscessus; Nontuberculous Mycobacteria; Pyrimidines; Pyrroles
PubMed: 36005813
DOI: 10.1128/aac.00669-22 -
Journal of Molecular Biology Dec 2018DNA gyrase, essential for DNA replication and transcription, has traditionally been studied in vivo by treatments that inhibit the enzyme activity. Due to its...
DNA gyrase, essential for DNA replication and transcription, has traditionally been studied in vivo by treatments that inhibit the enzyme activity. Due to its indispensable function, gyrA and gyrB deletions cannot be generated. The coumarin inhibitors of gyrase induce the supercoiling-sensitive gyrase promoter by a mechanism termed relaxation-stimulated transcription. Hence, to study the effect of sustained reduction in gyrase levels, a conditional-knockdown strain was generated in Mycobacterium smegmatis such that gyrase expression was controlled by a supercoiling non-responsive regulatory circuit. Decreasing intracellular gyrase protein levels beyond 50% affected cell growth. Reduced gyrase levels in the reprogrammed gyr operon caused chromosome relaxation, diffuse nucleoid structure, cell elongation, and altered gene expression. The key cell division protein, ftsZ, was severely reduced in the elongated cells, indicating a link between gyrase and cell division. Low levels of gyrase resulted in low compensatory expression of topoisomerase I and elevated expression of topology modulators hupB and lsr2. Altered supercoiling due to gyrase depletion caused corresponding changes in the RNA polymerase density on transcription units leading to their altered transcription. The enhanced susceptibility of the knockdown strain to anti-tubercular drugs suggests its utility for screening new molecules that may act synergistically with gyrase inhibitors.
Topics: Bacterial Proteins; Cell Division; Coumarins; DNA Gyrase; Gene Deletion; Gene Expression Regulation, Bacterial; Gene Knockdown Techniques; Mycobacterium smegmatis; Operon; Topoisomerase II Inhibitors; Transcription, Genetic
PubMed: 30316784
DOI: 10.1016/j.jmb.2018.10.001 -
Bioorganic Chemistry Dec 2022Antimicrobial resistance (AMR) is one of the critical challenges that have been encountered over the past years. On the other hand, bacterial DNA gyrase is regarded as...
Structure based design and synthesis of 3-(7-nitro-3-oxo-3,4-dihydroquinoxalin-2-yl)propanehydrazide derivatives as novel bacterial DNA-gyrase inhibitors: In-vitro, In-vivo, In-silico and SAR studies.
Antimicrobial resistance (AMR) is one of the critical challenges that have been encountered over the past years. On the other hand, bacterial DNA gyrase is regarded as one of the most outstanding biological targets that quinolones can extensively inhibit, improving AMR. Hence, a novel series of 3-(7-nitro-3-oxo-3,4-dihydroquinoxalin-2-yl)propanehydrazide derivatives (3-6j) were designed and synthesized employing the quinoxaline-2-one scaffold and relying on the pharmacophoric features experienced by the quinolone antibiotic; ciprofloxacin. The antibacterial activity of the synthesized compounds was assessed via in-vitro approaches using eight different Gram-positive and Gram-negative bacterial species. Most of the synthesized compounds revealed eligible antibacterial activities. In particular, compounds 6d and 6e displayed promising antibacterial activity among the investigated compounds. For example, compounds 6d and 6e displayed MIC values of 9.40 and 9.00 µM, respectively, regarding S. aureus, and 4.70 and 4.50 µM, respectively, regarding S. pneumonia in comparison to ciprofloxacin (12.07 µM). The cytotoxicity of compounds 6d and 6e were performed on normal human WI-38 cell lines with IC50 values of 288.69 and 227.64 μM, respectively assuring their safety and selectivity. Besides, DNA gyrase inhibition assay of compounds 6d and 6e was carried out in comparison to ciprofloxacin, and interestingly, compounds 6d and 6e disclosed promising IC50 values of 0.242 and 0.177 μM, respectively, whereas ciprofloxacin displayed an IC50 value of 0.768 μM, assuring the proposed mechanism of action for the afforded compounds. Consequently, compounds 6d and 6e were further assessed via in-vivo approaches by evaluating blood counts, liver and kidney functions, and histopathological examination. Both compounds were found to be safer on the liver and kidney than the reference ciprofloxacin. Moreover, in-silico molecular docking studies were established and revealed reasonable binding affinities for all afforded compounds, particularly compound 6d which exhibited a binding score of -7.51 kcal/mol, surpassing the reference ciprofloxacin (-7.29 kcal/mol) with better anticipated stability at the DNA gyrase binding pocket. Moreover, ADME studies were conducted, disclosing an eligible bioavailability score of >0.55 for all afforded compounds, and reasonable GIT absorption without passing the blood brain barrier was attained for most investigated compounds, ensuring their efficacy and safety. Lastly, a structure activity relationship study for the synthesized compounds was established and unveiled that not only the main pharmacophores required for DNA gyrase inhibition are enough for exerting promising antimicrobial activities, but also derivatization with diverse aryl/hetero aryl aldehydes is essential for their enhanced antimicrobial potential.
Topics: Humans; Anti-Bacterial Agents; Bacteria; Ciprofloxacin; DNA Gyrase; DNA, Bacterial; Microbial Sensitivity Tests; Molecular Docking Simulation; Molecular Structure; Quinolones; Staphylococcus aureus; Structure-Activity Relationship; Topoisomerase II Inhibitors
PubMed: 36215786
DOI: 10.1016/j.bioorg.2022.106186 -
Cell Reports Aug 2021The bacterial DNA gyrase complex (GyrA/GyrB) plays a crucial role during DNA replication and serves as a target for multiple antibiotics, including the fluoroquinolones....
The bacterial DNA gyrase complex (GyrA/GyrB) plays a crucial role during DNA replication and serves as a target for multiple antibiotics, including the fluoroquinolones. Despite it being a valuable antibiotics target, resistance emergence by pathogens including Pseudomonas aeruginosa are proving problematic. Here, we describe Igy, a peptide inhibitor of gyrase, encoded by Pseudomonas bacteriophage LUZ24 and other members of the Bruynoghevirus genus. Igy (5.6 kDa) inhibits in vitro gyrase activity and interacts with the P. aeruginosa GyrB subunit, possibly by DNA mimicry, as indicated by a de novo model of the peptide and mutagenesis. In vivo, overproduction of Igy blocks DNA replication and leads to cell death also in fluoroquinolone-resistant bacterial isolates. These data highlight the potential of discovering phage-inspired leads for antibiotics development, supported by co-evolution, as Igy may serve as a scaffold for small molecule mimicry to target the DNA gyrase complex, without cross-resistance to existing molecules.
Topics: DNA Gyrase; DNA Replication; DNA, Bacterial; Podoviridae; Pseudomonas Phages; Pseudomonas aeruginosa; Topoisomerase II Inhibitors; Viral Proteins
PubMed: 34433028
DOI: 10.1016/j.celrep.2021.109567 -
Chemical Biology & Drug Design Feb 2023This study aimed to synthesize new potent quinoline derivatives based on hydrazone moieties and evaluate their antimicrobial activity. The newly synthesized...
Development and radiosterilization of new hydrazono-quinoline hybrids as DNA gyrase and topoisomerase IV inhibitors: Antimicrobial and hemolytic activities against uropathogenic isolates with molecular docking study.
This study aimed to synthesize new potent quinoline derivatives based on hydrazone moieties and evaluate their antimicrobial activity. The newly synthesized hydrazono-quinoline derivatives 2, 5a, 9, and 10b showed the highest antimicrobial activity with MIC values ≤1.0 μg/ml against bacteria and ≤8.0 μg/ml against the fungi. Further, these derivatives exhibited bactericidal and fungicidal effects with MBC/MIC and MFC/MIC ratio ≤4. Surprisingly, the most active compounds displayed good inhibition to biofilm formation with MBEC values ranging between (40.0 ± 10.0 - 230.0 ± 31.0) and (67.0 ± 24.0 - 347.0 ± 15.0) μg/ml against Staphylococcus aureus and Pseudomonas aeruginosa, respectively. The hemolytic assays confirmed that the hydrazono-quinoline derivatives are non-toxic with low % lysis values ranging from 4.62% to 14.4% at a 1.0 mg/ml concentration. Besides, compound 5a exhibited the lowest hemolytic activity value of ~4.62%. Furthermore, the study suggests that the hydrazono-quinoline analogs exert their antibacterial activity as dual inhibitors for DNA gyrase and DNA topoisomerase IV enzymes with IC values ranging between (4.56 ± 0.3 - 21.67 ± 0.45) and (6.77 ± 0.4 - 20.41 ± 0.32) μM, respectively. Additionally, the recent work advocated that compound 5a showed the reference SAL at the ɣ-radiation dose of 10.0 kGy in the sterilization process without affecting its chemical structure. Finally, the in silico drug-likeness, toxicity properties, and molecular docking simulation were performed. Besides, the result exhibited good oral-bioavailability, lower toxicity prediction, and lower binding energy with good binding mode rather than the positive control.
Topics: Molecular Docking Simulation; DNA Gyrase; DNA Topoisomerase IV; Topoisomerase II Inhibitors; Anti-Infective Agents; Anti-Bacterial Agents; Microbial Sensitivity Tests; Structure-Activity Relationship; Molecular Structure
PubMed: 36305722
DOI: 10.1111/cbdd.14154 -
Journal of Inorganic Biochemistry Aug 2021An oxovanadium(IV) - curcumin based complex, viz. [VO(cur)(2,2´-bipy)(HO)] where cur is curcumin and bipy is bipyridine, previously synthesized, has been studied for...
Unraveling the binding mechanism of an Oxovanadium(IV) - Curcumin complex on albumin, DNA and DNA gyrase by in vitro and in silico studies and evaluation of its hemocompatibility.
An oxovanadium(IV) - curcumin based complex, viz. [VO(cur)(2,2´-bipy)(HO)] where cur is curcumin and bipy is bipyridine, previously synthesized, has been studied for interaction with albumin and DNA. Fluorescence emission spectroscopy was used to evaluate the interaction of the complex with bovine serum albumin (BSA) and the BSA-binding constant (K) was calculated to be 2.56 x 10 M, whereas a single great-affinity binding site was revealed. Moreover, the hemocompatibility test demonstrated that the complex presented low hemolytic fraction (mostly below 1%), in all concentrations tested (0-250 μΜ of complex, 5% DMSO) assuring a safe application in interaction with blood. The binding of the complex to DNA was also investigated using absorption, fluorescence, and viscometry methods indicating a binding through a minor groove mode. From competitive studies with ethidium bromide the apparent binding constant value to DNA was estimated to be 4.82 x 10 M. Stern-Volmer quenching phenomenon gave a Κ constant [1.92 (± 0.05) x 10 M] and k constant [8.33 (± 0.2) x 10 Ms]. Molecular docking simulations on the crystal structure of BSA, calf thymus DNA, and DNA gyrase, as well as pharmacophore analysis for BSA target, were also employed to study in silico the ability of [VO(cur)(2,2´-bipy)(HO)] to bind to these target bio-macromolecules and explain the observed in vitro activity.
Topics: Animals; Binding Sites; Cattle; Coordination Complexes; Curcumin; DNA; DNA Gyrase; Escherichia coli; Escherichia coli Proteins; Hemolysis; Humans; Ligands; Molecular Docking Simulation; Protein Binding; Serum Albumin, Bovine; Vanadium; Viscosity
PubMed: 33975249
DOI: 10.1016/j.jinorgbio.2021.111402