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Scientific Reports Jan 2023Flavonoids are plant-produced secondary metabolites that are found ubiquitously. We have previously reported that apigenin, a class of flavonoid, has unique...
Flavonoids are plant-produced secondary metabolites that are found ubiquitously. We have previously reported that apigenin, a class of flavonoid, has unique antimicrobial activity against Staphylococcus aureus (S. aureus), one of the major human pathogens. Apigenin inhibited fluoroquinolone-resistant S. aureus with DNA gyrase harboring the quinolone-resistant S84L mutation but did not inhibit wild-type DNA gyrase. In this study, we describe five flavonoids, quercetin, luteolin, kaempferol, baicalein, and commercially available CID12261165, that show similar antimicrobial activity against fluoroquinolone-resistant S. aureus. Among them, CID12261165 was the most effective with MIC values of ≤ 4 mg/L against quinolone-resistant S. aureus strains. In vitro DNA cleavage and supercoiling assays demonstrated inhibitory activity of CID12261165 against mutated DNA gyrase, whereas activity against wild-type DNA gyrase was not observed. CID12261165 also inhibited quinolone-resistant Enterococci with an MIC value of 8 mg/L. While fluoroquinolone-resistant amino acid replacements can improve the fitness of bacterial cells, it is unknown why quinolone-susceptible S. aureus strains were predominant before the introduction of fluoroquinolone. The present study discusses the current discrepancies in the interpretation of antimicrobial activities of flavonoids, as well as the possible reasons for the preservation of wild-type DNA gyrase wherein the environmental flavonoids cannot be ignored.
Topics: Anti-Bacterial Agents; Apigenin; DNA Gyrase; Flavonoids; Fluoroquinolones; Staphylococcus aureus; Drug Resistance, Bacterial
PubMed: 36720958
DOI: 10.1038/s41598-023-28859-8 -
Frontiers in Bioengineering and... 2021Antibacterial resistance (ABR) is a major life-threatening problem worldwide. Rampant dissemination of ABR always exemplified the need for the discovery of novel...
Antibacterial resistance (ABR) is a major life-threatening problem worldwide. Rampant dissemination of ABR always exemplified the need for the discovery of novel compounds. However, to circumvent the disease, a molecular target is required, which will lead to the death of the bacteria when acted upon by a compound. One group of enzymes that have proved to be an effective target for druggable candidates is bacterial DNA topoisomerases (DNA gyrase and ParE). In our present work, phenylacetamide and benzohydrazides derivatives were screened for their antibacterial activity against a selected panel of pathogens. The tested compounds displayed significant antibacterial activity with MIC values ranging from 0.64 to 5.65 μg/mL. Amongst 29 title compounds, compounds 5 and 21 exhibited more potent and selective inhibitory activity against with MIC values at 0.64 and 0.67 μg/mL, respectively, and MBC at onefold MIC. Furthermore, compounds exhibited a post-antibiotic effect of 2 h at 1× MIC in comparison to ciprofloxacin and gentamicin. These compounds also demonstrated the concentration-dependent bactericidal activity against and synergized with FDA-approved drugs. The compounds are screened for their enzyme inhibitory activity against ParE, whose IC values range from 0.27 to 2.80 μg/mL. Gratifyingly, compounds, namely 8 and 25 belonging to the phenylacetamide series, were found to inhibit ParE enzyme with IC values of 0.27 and 0.28 μg/mL, respectively. In addition, compounds were benign to Vero cells and displayed a promising selectivity index (169.0629-951.7240). Moreover, compounds 1, 7, 8, 21, 24, and 25 (IC: <1 and Selectivity index: >200) exhibited potent activity in reducing the biofilm in comparison with ciprofloxacin, erythromycin, and ampicillin. These astonishing results suggest the potential utilization of phenylacetamide and benzohydrazides derivatives as promising ParE inhibitors for treating bacterial infections.
PubMed: 34222214
DOI: 10.3389/fbioe.2021.669728 -
ACS Omega Nov 2019DNA topoisomerases are essential enzymes for all living organisms and important targets for anticancer drugs and antibiotics. Although DNA topoisomerases have been...
DNA topoisomerases are essential enzymes for all living organisms and important targets for anticancer drugs and antibiotics. Although DNA topoisomerases have been studied extensively, steady-state kinetics has not been systematically investigated because of the lack of an appropriate assay. Previously, we demonstrated that newly synthesized, fluorescently labeled plasmids pAB1_FL905 and pAB1_FL924 can be used to study DNA topoisomerase-catalyzed reactions by fluorescence resonance energy transfer (FRET) or supercoiling-dependent fluorescence quenching (SDFQ). With the FRET or SDFQ method, we performed steady-state kinetic studies for six different DNA topoisomerases including two type IA enzymes ( and DNA topoisomerase I), two type IB enzymes (human and variola DNA topoisomerase I), and two type IIA enzymes ( DNA gyrase and human DNA topoisomerase IIα). Our results show that all DNA topoisomerases follow the classical Michaelis-Menten kinetics and have unique steady-state kinetic parameters, , , and . We found that for all topoisomerases are rather low and that such low values may stem from the tight binding of topoisomerases to DNA. Additionally, we confirmed that novobiocin is a competitive inhibitor for adenosine 5'-triphosphate binding to DNA gyrase, demonstrating the utility of our assay for studying topoisomerase inhibitors.
PubMed: 31720544
DOI: 10.1021/acsomega.9b02676 -
Applied and Environmental Microbiology Jul 2021Spread of biosolids-borne antibiotic resistance is a growing public and environmental health concern. Herein, we conducted incubation experiments involving biosolids,...
Spread of biosolids-borne antibiotic resistance is a growing public and environmental health concern. Herein, we conducted incubation experiments involving biosolids, which are byproducts of sewage treatment processes, and biosolids-amended soil. Quantitative reverse transcription-PCR (RT-qPCR) was employed to assess responses of select antibiotic resistance genes (ARGs) and mobile elements to environmentally relevant concentrations of two biosolids-borne antibiotics, azithromycin (AZ) and ciprofloxacin (CIP). Additionally, we examined sequence distribution of (encoding DNA gyrase; site of action of CIP) to assess potential shifts in genotype. Increasing antibiotic concentrations generally increased the transcriptional activities of (encoding CIP resistance) and and (encoding AZ resistance). The transcriptional activity of , a marker of class 1 integrons, was unaffected by CIP or AZ concentrations, but biosolids amendment increased activity in the soil by 4 to 5 times, which persisted throughout incubation. While the dominant sequences found herein were unrelated to known CIP-resistant genotypes, the increasing CIP concentrations significantly decreased the diversity of genes encoding the DNA gyrase A subunit, suggesting changes in microbial community structures. This study suggests that biosolids harbor transcriptionally active ARGs and mobile elements that could survive and spread in biosolids-amended soils. However, more research is warranted to investigate these trends under field conditions. Although previous studies have indicated that biosolids may be important spreaders of antibiotics and antibiotic resistance genes (ARGs) in environments, the potential activities of ARGs or their responses to environmental parameters have been understudied. This study highlights that certain biosolids-borne antibiotics can induce transcriptional activities of ARGs and mobile genetic elements in biosolids and biosolids-amended soil, even when present at environmentally relevant concentrations. Furthermore, these antibiotics can alter the structure of microbial populations expressing ARGs. Our findings indicate the bioavailability of the antibiotics in biosolids and provide evidence that biosolids can promote the activities and dissemination of ARGs and mobile genes in biosolids and soils that receive contaminated biosolids, thus, underscoring the importance of investigating anthropogenically induced antibiotic resistance in the environment under real-world scenarios.
Topics: Anti-Bacterial Agents; Azithromycin; Bacteria; Bacterial Proteins; Biosolids; Ciprofloxacin; Drug Resistance, Bacterial; Interspersed Repetitive Sequences; Soil; Soil Microbiology; Soil Pollutants
PubMed: 34085858
DOI: 10.1128/AEM.00373-21 -
ACS Infectious Diseases Apr 2024Antimicrobial resistance is a global threat to human health. Therefore, efforts have been made to develop new antibacterial agents that address this critical medical...
Antimicrobial resistance is a global threat to human health. Therefore, efforts have been made to develop new antibacterial agents that address this critical medical issue. Gepotidacin is a novel, bactericidal, first-in-class triazaacenaphthylene antibacterial in clinical development. Recently, phase III clinical trials for gepotidacin treatment of uncomplicated urinary tract infections caused by uropathogens, including , were stopped for demonstrated efficacy. Because of the clinical promise of gepotidacin, it is important to understand how the compound interacts with its cellular targets, gyrase and topoisomerase IV, from . Consequently, we determined how gyrase and topoisomerase IV mutations in amino acid residues that are involved in gepotidacin interactions affect the susceptibility of cells to the compound and characterized the effects of gepotidacin on the activities of purified wild-type and mutant gyrase and topoisomerase IV. Gepotidacin displayed well-balanced dual-targeting of gyrase and topoisomerase IV in cells, which was reflected in a similar inhibition of the catalytic activities of these enzymes by the compound. Gepotidacin induced gyrase/topoisomerase IV-mediated single-stranded, but not double-stranded, DNA breaks. Mutations in GyrA and ParC amino acid residues that interact with gepotidacin altered the activity of the compound against the enzymes and, when present in both gyrase and topoisomerase IV, reduced the antibacterial activity of gepotidacin against this mutant strain. Our studies provide insights regarding the well-balanced dual-targeting of gyrase and topoisomerase IV by gepotidacin in .
Topics: Acenaphthenes; Amino Acids; Anti-Bacterial Agents; DNA Gyrase; DNA Topoisomerase IV; Escherichia coli; Heterocyclic Compounds, 3-Ring
PubMed: 38606465
DOI: 10.1021/acsinfecdis.3c00346 -
Life (Basel, Switzerland) Feb 2023sp. AKA109 is a producer of akazaoxime and A-76356, whereas sp. AKA38 is that of levantilide C. We aimed to clarify their taxonomic positions and identify biosynthetic...
sp. AKA109 is a producer of akazaoxime and A-76356, whereas sp. AKA38 is that of levantilide C. We aimed to clarify their taxonomic positions and identify biosynthetic gene clusters (BGCs) of these compounds. In 16S rRNA gene and DNA gyrase subunit B gene () sequence analyses, strains AKA109 and AKA38 were the most closely related to MMS20-R2-29 and HKI0641, respectively. Although sp. AKA109 was identified as by the sequence similarity and DNA-DNA relatedness based on whole genome sequences, sp. AKA38 was classified to a new genomospecies. AKA109 harbored six type-I polyketide synthase (PKS), one type-II PKS, one type-III PKS, three non-ribosomal peptide synthetase (NRPS) and three hybrid PKS/NRPS gene clusters, among which the BGC of akazaoxime and A-76356 was identified. These gene clusters are conserved in MMS20-R2-29 sp. AKA38 harbored two type-I PKS, one of which was responsible for levantilide C, one type-II PKS, one type-III PKS, two NRPS and five hybrid PKS/NRPS gene clusters. We predicted products derived from these gene clusters through bioinformatic analyses. Consequently, these two strains are revealed to be promising sources for diverse non-ribosomal peptide and polyketide compounds.
PubMed: 36836900
DOI: 10.3390/life13020542 -
Bioorganic & Medicinal Chemistry May 2020Fluoroquinolones are a class of antibacterial agents used clinically to treat a wide array of bacterial infections and target bacterial type-II topoisomerases (DNA...
Fluoroquinolones are a class of antibacterial agents used clinically to treat a wide array of bacterial infections and target bacterial type-II topoisomerases (DNA gyrase and topoisomerase IV). Fluoroquinolones, however potent, are susceptible to bacterial resistance with prolonged use, which limits their use in the clinic. Quinazoline-2,4-diones also target bacterial type-II topoisomerases and are not susceptible to bacterial resistance similar to fluoroquinolones, however, their potency pales in comparison to fluoroquinolones. To meet the increasing demand for antibacterial development, nine modified quinazoline-2,4-diones were developed to probe quinazoline-2,4-dione structure modification for possible new binding contacts with the bacterial type-II topoisomerase, DNA gyrase. Evaluation of compounds for inhibition of the supercoiling activity of DNA gyrase revealed a novel ethyl 5,6-dihydropyrazolo[1,5-c]quinazoline-1-carboxylate derivative as a modest inhibitor of DNA gyrase, having an IC of 3.5 μM. However, this ethyl 5,6-dihydropyrazolo[1,5-c]quinazoline-1-carboxylate does not trap the catalytic intermediate like fluoroquinolones or typical quinazoline-2,4-diones do. Thus, the ethyl 5,6-dihydropyrazolo[1,5-c]quinazoline-1-carboxylate derivative discovered in this work acts as a catalytic inhibitor of DNA gyrase and therefore represents a new structural type of catalytic inhibitor of DNA gyrase.
Topics: Biocatalysis; DNA Gyrase; Dose-Response Relationship, Drug; Escherichia coli; Molecular Structure; Structure-Activity Relationship; Topoisomerase II Inhibitors
PubMed: 32234278
DOI: 10.1016/j.bmc.2020.115439 -
International Journal For Parasitology May 2021Apicomplexans are the causative agents of numerous important infectious diseases including malaria and toxoplasmosis. Most of them harbour a chloroplast-like organelle...
Apicomplexans are the causative agents of numerous important infectious diseases including malaria and toxoplasmosis. Most of them harbour a chloroplast-like organelle called the apicoplast that is essential for the parasites' metabolism and survival. While most apicoplast proteins are nuclear encoded, the organelle also maintains its own genome, a 35 kb circle. In this study we used Toxoplasma gondii to identify and characterise essential proteins involved in apicoplast genome replication and to understand how apicoplast genome segregation unfolds over time. We demonstrated that the DNA replication enzymes Prex, DNA gyrase and DNA single stranded binding protein localise to the apicoplast. We show in knockdown experiments that apicoplast DNA Gyrase A and B, and Prex are required for apicoplast genome replication and growth of the parasite. Analysis of apicoplast genome replication by structured illumination microscopy in T. gondii tachyzoites showed that apicoplast nucleoid division and segregation initiate at the beginning of S phase and conclude during mitosis. Thus, the replication and division of the apicoplast nucleoid is highly coordinated with nuclear genome replication and mitosis. Our observations highlight essential components of apicoplast genome maintenance and shed light on the timing of this process in the context of the overall parasite cell cycle.
Topics: Apicoplasts; Cell Division; DNA Gyrase; DNA-Directed DNA Polymerase; Humans; Toxoplasma; Toxoplasmosis
PubMed: 33581138
DOI: 10.1016/j.ijpara.2020.11.004 -
Journal of Enzyme Inhibition and... Dec 2022Emerging drug resistance is generating an urgent need for novel and effective antibiotics. A promising target that has not yet been addressed by approved antibiotics is...
Emerging drug resistance is generating an urgent need for novel and effective antibiotics. A promising target that has not yet been addressed by approved antibiotics is the bacterial DNA gyrase subunit B (GyrB), and GyrB inhibitors could be effective against drug-resistant bacteria, such as methicillin-resistant (MRSA). Here, we used the 4-hydroxy-2-quinolone fragment to search the Specs database of purchasable compounds for potential inhibitors of GyrB and identified or , as a novel and potent inhibitor of the target protein (IC: 1.21 µM). Structural modification was used to further identify two more potent GyrB inhibitors: (IC: 0.31 µM) and (IC: 0.28 µM). Additional experiments indicated that compound is more potent than the others in terms of antibacterial activity against MRSA (MICs: 4-8 µg/mL), non-toxic to HUVEC and HepG2 (CC: approximately 50 µM), and metabolically stable (t: > 372.8 min for plasma; 24.5 min for liver microsomes). In summary, this study showed that the discovered N-quinazolinone-4-hydroxy-2-quinolone-3-carboxamides are novel GyrB-targeted antibacterial agents; compound is promising for further development.
Topics: DNA Gyrase; Anti-Bacterial Agents; Topoisomerase II Inhibitors; Methicillin-Resistant Staphylococcus aureus; Quinazolinones; DNA, Bacterial; Microbial Sensitivity Tests; Bacteria
PubMed: 36278813
DOI: 10.1080/14756366.2022.2084088 -
Antimicrobial Agents and Chemotherapy Mar 2022Antibiotics are the current standard-of-care treatment for uncomplicated urinary tract infections (uUTIs). However, increasing rates of bacterial antibiotic resistance... (Review)
Review
Antibiotics are the current standard-of-care treatment for uncomplicated urinary tract infections (uUTIs). However, increasing rates of bacterial antibiotic resistance necessitate novel therapeutic options. Gepotidacin is a first-in-class triazaacenaphthylene antibiotic that selectively inhibits bacterial DNA replication by interaction with the bacterial subunits of DNA gyrase (GyrA) and topoisomerase IV (ParC). Gepotidacin is currently in clinical development for the treatment of uUTIs and other infections. In this article, we review data for gepotidacin from nonclinical studies, including activity, animal efficacy, and pharmacokinetic (PK) and pharmacokinetic/pharmacodynamic (PK/PD) models that informed dose selection for phase III clinical evaluation of gepotidacin. Based on this translational package of data, a gepotidacin 1,500-mg oral dose twice daily for 5 days was selected for two ongoing, randomized, multicenter, parallel-group, double-blind, double-dummy, active-comparator phase III clinical studies evaluating the safety and efficacy of gepotidacin in adolescent and adult female participants with uUTIs (ClinicalTrials.gov identifiers NCT04020341 and NCT04187144).
Topics: Acenaphthenes; Adolescent; Animals; Anti-Bacterial Agents; Clinical Trials, Phase III as Topic; Female; Heterocyclic Compounds, 3-Ring; Humans; Microbial Sensitivity Tests; Multicenter Studies as Topic; Randomized Controlled Trials as Topic; Urinary Tract Infections
PubMed: 34978887
DOI: 10.1128/AAC.01492-21