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Antibiotics (Basel, Switzerland) Jul 2022The free-living amoeba is responsible for the central nervous infection granulomatous amoebic encephalitis and sight-threatening infection . Moreover, no effective...
The free-living amoeba is responsible for the central nervous infection granulomatous amoebic encephalitis and sight-threatening infection . Moreover, no effective treatment is currently present, and a combination drug therapy is used. In this study, twelve DNA minor groove binders (MGBs) were synthesized and tested for their antiamoebic activity via amoebicidal, encystation, excystation, and cytopathogenicity assays. It was found that the compounds MGB3, MGB6, MGB22, MGB24, and MGB16 significantly reduce amoeba viability to 76.20%, 59.45%, 66.5%, 39.32%, and 43.21%, respectively, in amoebicidal assays. Moreover, the compounds MGB6, MGB20, MGB22, MGB28, MGB30, MGB32, and MGB16 significantly inhibit cysts, leading to the development of only 46.3%, 39%, 30.3%, 29.6%, 27.8%, 41.5%, and 45.6% cysts. Additionally, the compounds MGB3, MGB4, MGB6, MGB22, MGB24, MGB28, MGB32, and MGB16 significantly reduce the re-emergence of cysts to trophozoites, with viable trophozoites being only 64.3%, 47.3%, 41.4%, 52.9%, 55.4%, 40.6%, 62.1%, and 51.7%. Moreover, the compounds MGB3, MGB4, and MGB6 exhibited the greatest reduction in amoeba-mediated host-cell death, with cell death reduced to 41.5%, 49.4%, and 49.5%. With the following determined, future in vivo studies can be carried out to understand the effect of the compounds on animal models such as mice.
PubMed: 35884189
DOI: 10.3390/antibiotics11070935 -
Applied and Environmental Microbiology Nov 2021The production of specialized metabolites by bacteria is usually temporally regulated. This regulation is complex and frequently involves both global and...
The production of specialized metabolites by bacteria is usually temporally regulated. This regulation is complex and frequently involves both global and pathway-specific mechanisms. Streptomyces ambofaciens ATCC23877 produces several specialized metabolites, including spiramycins, stambomycins, kinamycins and congocidine. The production of the first three molecules has been shown to be controlled by one or several cluster-situated transcriptional regulators. However, nothing is known regarding the regulation of congocidine biosynthesis. Congocidine (netropsin) belongs to the family of pyrrolamide metabolites, which also includes distamycin and anthelvencins. Most pyrrolamides bind into the minor groove of DNA, specifically in A/T-rich regions, which gives them numerous biological activities, such as antimicrobial and antitumoral activities. We previously reported the characterization of the pyrrolamide biosynthetic gene clusters of congocidine () in S. ambofaciens ATCC23877, distamycin () in Streptomyces netropsis DSM40846, and anthelvencins () in Streptomyces venezuelae ATCC14583. The three gene clusters contain a gene encoding a putative transcriptional regulator, , , and respectively. Cgc1, Dst1, and Ant1 present a high percentage of amino acid sequence similarity. We demonstrate here that Cgc1, an atypical orphan response regulator, activates the transcription of all genes in the stationary phase of S. ambofaciens growth. We also show that the cluster is constituted of eight main transcriptional units. Finally, we show that congocidine induces the expression of the transcriptional regulator Cgc1 and of the operon containing the resistance genes ( and , coding for an ABC transporter), and propose a model for the transcriptional regulation of the gene cluster. Understanding the mechanisms of regulation of specialized metabolite production can have important implications both at the level of specialized metabolism study (expression of silent gene clusters) and at the biotechnological level (increase of the production of a metabolite of interest). We report here a study on the regulation of the biosynthesis of a metabolite from the pyrrolamide family, congocidine. We show that congocidine biosynthesis and resistance are controlled by Cgc1, a cluster-situated regulator. As the gene clusters directing the biosynthesis of the pyrrolamides distamycin and anthelvencin encode a homolog of Cgc1, our findings may be relevant for the biosynthesis of other pyrrolamides. In addition, our results reveal a new type of feed-forward induction mechanism, in which congocidine induces its own biosynthesis through the induction of the transcription of .
Topics: Distamycins; Gene Expression Regulation, Bacterial; Genes, Bacterial; Multigene Family; Netropsin; Streptomyces
PubMed: 34586912
DOI: 10.1128/AEM.01380-21 -
Molecules (Basel, Switzerland) Aug 2021The recognition of specific DNA sequences in processes such as transcription is associated with a cooperative binding of proteins. Some transcription regulation...
The recognition of specific DNA sequences in processes such as transcription is associated with a cooperative binding of proteins. Some transcription regulation mechanisms involve additional proteins that can influence the binding cooperativity by acting as corepressors or coactivators. In a conditional cooperativity mechanism, the same protein can induce binding cooperativity at one concentration and inhibit it at another. Here, we use calorimetric (ITC) and spectroscopic (UV, CD) experiments to show that such conditional cooperativity can also be achieved by the small DNA-directed oligopeptides distamycin and netropsin. Using a global thermodynamic analysis of the observed binding and (un)folding processes, we calculate the phase diagrams for this system, which show that distamycin binding cooperativity is more pronounced at lower temperatures and can be first induced and then reduced by increasing the netropsin or/and Na+ ion concentration. A molecular interpretation of this phenomenon is suggested.
Topics: DNA; Distamycins; Netropsin; Oligopeptides; Protein Binding; Sodium; Thermodynamics; Transcription, Genetic
PubMed: 34500619
DOI: 10.3390/molecules26175188 -
In Silico Pharmacology 2021Severe Acute Respiratory Syndrome coronavirus (SARS-CoV), Middle East Respiratory Syndrome coronavirus (MERS-CoV) and the novel SARS-CoV-2 evade the host innate...
UNLABELLED
Severe Acute Respiratory Syndrome coronavirus (SARS-CoV), Middle East Respiratory Syndrome coronavirus (MERS-CoV) and the novel SARS-CoV-2 evade the host innate immunity, and subsequently the adaptive immune response, employing one protease called Papain-like protease (PLpro). The PLpro and the 3CL main protease are responsible for the cleavage of the polyproteins encoded by the + sense RNA genome of the virus to produce several non-structured proteins (NSPs). However, the PLpro also performs deubiquitination and deISGylation of host proteins and signaling molecules, and thus antagonize the host innate immune response, since ubiquitination and ISGylation are critical processes which invoke host's antiviral immune responses. Thus, to maintain host antiviral defense, inhibition of the PLpro is the primary therapeutic strategy. Furthermore, inhibition of the enzyme prevents replication of the virus. The present study employs molecular modeling approaches to determine potential of different approved and repurposed drugs and other compounds as inhibitors of the SARS-CoV-2 PLpro. The results of the study demonstrated that drugs like Stallimycin, and known protease inhibitors including Telaprevir, Grazoprevir and Boceprevir, were highly potent in inhibiting the enzyme. In addition, several plant-derived polyphenols, including Corylifol A and Kazinol J, were found to be potent inhibitors. Based on the findings, we suggest that clinical trials be initiated with these inhibitors. So far, PLpro inhibition has been given less attention as a strategy to contain COVID-19 pandemic, and thus the present study is of high significance and has therapeutic implications in containing the pandemic.
SUPPLEMENTARY INFORMATION
The online version contains supplementary material available at 10.1007/s40203-021-00085-y.
PubMed: 33842190
DOI: 10.1007/s40203-021-00085-y -
Archives of Biochemistry and Biophysics Apr 2021Human telomerase that activates within cancer cells has a telomeric sequence at the 3' end. Each factor that stabilizes the G-quadruplex in guanine-rich telomeric...
Human telomerase that activates within cancer cells has a telomeric sequence at the 3' end. Each factor that stabilizes the G-quadruplex in guanine-rich telomeric sequences can inhibit the regular telomerase activity. Therefore, the telomeric G-quadruplex is known as a promising target in cancer treatment. In this work, we studied the binding of positively charged distamycin A and its uncharged derivative to the G-quadruplex in a solution environment by Molecular Dynamics (MD) simulation. The binding mechanism and subtle conformational changes were investigated as a result of the ligand attachment. Moreover, binding free energy and clustering analysis describe the stability and flexibility of G-quadruplexes upon ligand binding. Structural analyses displayed that the favorable binding of both ligands imposes significant stability and rigidity in G-quadruplex conformation compared to free G-quadruplex, especially charged distamycin. Hydration pattern and ion distribution were different for free G-quadruplex and both of the ligand complexes. Energy decomposition reveals the electrostatic effect on the stability of G-quadruplex. The radial distribution function displayed the solvent shell and ion moving away from the groove. The hydrogen bond played an essential role in the binding of both ligands, especially for the charged derivative. van der Waals interaction is the only factor that is more important in binding uncharged distamycin into G-quadruplex than the charged one. The calculated ΔG showed the stability of both ligands within grooves and good agreement with the experimental binding free energy data. Finally, the results suggest that ligand modification improves the binding mode toward stabilizing G-quadruplexes.
Topics: Antineoplastic Agents; Distamycins; G-Quadruplexes; Humans; Molecular Dynamics Simulation; Telomere
PubMed: 33607110
DOI: 10.1016/j.abb.2021.108797 -
Chemistry (Weinheim An Der Bergstrasse,... May 2021The term "privileged structure" refers to a single molecular substructure or scaffold that can serve as a starting point for high-affinity ligands for more than one... (Review)
Review
The term "privileged structure" refers to a single molecular substructure or scaffold that can serve as a starting point for high-affinity ligands for more than one receptor type. In this report, a hitherto overlooked group of privileged substructures is addressed, namely aromatic oligoamides, for which there are natural models in the form of cystobactamids, albicidin, distamycin A, netropsin, and others. The aromatic and heteroaromatic core, together with a flexible selection of substituents, form conformationally well-defined scaffolds capable of specifically binding to conformationally well-defined regions of biomacromolecules such as helices in proteins or DNA often by acting as helices mimics themselves. As such, these aromatic oligoamides have already been employed to inhibit protein-protein and nucleic acid-protein interactions. This article is the first to bring together the scattered knowledge about aromatic oligoamides in connection with biomedical applications.
Topics: DNA; Ligands; Protein Structure, Secondary; Proteins
PubMed: 33481284
DOI: 10.1002/chem.202005086 -
European Journal of Medicinal Chemistry Jun 2020We have designed and synthesized anthraquinone containing compounds which have oligopyrrole side chains of varying lengths. These compounds stabilized the G-quadruplex...
Specific stabilization of promoter G-Quadruplex DNA by 2,6-disubstituted amidoanthracene-9,10-dione based dimeric distamycin analogues and their selective cancer cell cytotoxicity.
We have designed and synthesized anthraquinone containing compounds which have oligopyrrole side chains of varying lengths. These compounds stabilized the G-quadruplex DNA formed in the promoter regions of c-MYC oncogenes selectively over the duplex DNA. These observations were recorded using UV-vis spectroscopic titrations, fluorescence measurements and circular dichroism (CD) spectral titrations. The potency of the compounds to stabilize the G4 DNA has been shown from the thermal denaturation experiments. The compound interacts with c-MYC G-quadruplex DNA through stacking mode as obtained from ethidium bromide displacement assay, cyclic voltammetric titration, and docking experiments. Molecular modeling studies suggested that the stacking of the anthraquinone moiety over the G-tetrad of the G4 structures are responsible for the stability of such quadruplex secondary structure. Furthermore, polymerase stop assay also supported the formation of stable G4 structures in the presence of the above-mentioned compounds. The compounds have shown selective cancer cell (HeLa and HEK293T) cytotoxicity over normal cells (NIH3T3 and HDFa) under in vitro conditions as determined from MTT based cell viability assay. Apoptosis was found to be the mechanistic pathway underlying the cancer cell cytotoxicity as obtained from Annexin V-FITC and PI dual staining assay which was further substantiated by nuclear morphological changes as observed by AO/EB dual staining assay. Cellular morphological changes, as well as nuclear condensation and fragmentation upon treatment with these compounds, were observed under bright field and confocal microscopy.
Topics: Anthracenes; Antineoplastic Agents; DNA; Dimerization; Distamycins; Drug Design; G-Quadruplexes; Models, Molecular; Promoter Regions, Genetic; Proto-Oncogene Proteins c-myc
PubMed: 32302880
DOI: 10.1016/j.ejmech.2020.112202 -
ACS Chemical Biology Apr 2020Anthelvencins A and B are pyrrolamide metabolites produced by ATCC 14583 and 14585. Isolated in 1965, they were reported to exhibit anthelmintic and moderate...
Anthelvencins A and B are pyrrolamide metabolites produced by ATCC 14583 and 14585. Isolated in 1965, they were reported to exhibit anthelmintic and moderate antibacterial activities. In this study, we revise the structure of anthelvencin A and identify a third anthelvencin metabolite, bearing two -methylated pyrrole groups, which we named anthelvencin C. We sequenced the genome of ATCC 14583 and identified a gene cluster predicted to direct the biosynthesis of anthelvencins. Functional analysis of this gene cluster confirmed its involvement in anthelvencin biosynthesis and allowed us to propose a biosynthetic pathway for anthelvencins. In addition to a nonribosomal peptide synthetase (NRPS), the assembly of anthelvencins involves an enzyme from the ATP-grasp ligase family, Ant23. We propose that Ant23 uses a PCP-loaded 4-aminopyrrole-2-carboxylate as substrate. As observed for the biosynthesis of the other pyrrolamides congocidine (produced by ATCC 25877) and distamycin (produced by DSM 40846), the NRPS assembling anthelvencins is composed of stand-alone domains only. Such NRPSs, sometimes called type II NRPSs, are less studied than the classical multimodular NRPSs. Yet, they constitute an interesting model to study protein-protein interactions in NRPSs and are good candidates for combinatorial biosynthesis approaches.
Topics: Bacterial Proteins; Multigene Family; Peptide Synthases; Protein Domains; Pyrroles; Streptomyces
PubMed: 32129986
DOI: 10.1021/acschembio.9b00960 -
European Journal of Medicinal Chemistry Mar 2020Polyamides-based compounds related to the Streptomycetal distamycin and netropsin are potent cytostatic molecules that bind to AT-rich regions of the minor groove of the...
Polyamides-based compounds related to the Streptomycetal distamycin and netropsin are potent cytostatic molecules that bind to AT-rich regions of the minor groove of the DNA, hence interfering with DNA replication and transcription. Recently, derivatives belonging to this scaffold have been reported to halt the proliferation of deadly African trypanosomes by different and unrelated mechanisms. Here we describe the synthesis and preliminary characterization of the anti-trypanosomal mode of action of new potent and selective distamycin analogues. Two tri-heterocyclic derivatives containing a central N-methyl pyrrole ring (16 and 17) displayed high activity (EC < 20 nM) and selectivity (selectivity index >5000 with respect to mammalian macrophages) against the infective form of T. brucei. Both compounds caused cell cycle arrest by blocking the replication of the mitochondrial DNA but without affecting its integrity. This mode of action clearly differs from that reported for classical minor groove binder (MGB) drugs, which induce the degradation of the mitochondrial DNA. In line with this, in vitro assays suggest that 16 and 17 have a comparatively lower affinity for different template DNAs than the MGB drug diminazene. Therapeutic efficacy studies and stability assays suggest that the pharmacological properties of the hits should be optimized. The compounds can be rated as excellent scaffolds for the design of highly potent and selective anti-T. brucei agents.
Topics: Animals; Cell Cycle; Distamycins; Female; Macrophages; Mice; Mice, Inbred BALB C; Thiazoles; Trypanocidal Agents; Trypanosoma; Trypanosoma brucei brucei; Trypanosomiasis, African
PubMed: 31978782
DOI: 10.1016/j.ejmech.2020.112043 -
Journal of Computational Chemistry Apr 2020Alchemically derived free energies are artifacted when the perturbed moiety has a nonzero net charge. The source of the artifacts lies in the effective treatment of the...
Alchemically derived free energies are artifacted when the perturbed moiety has a nonzero net charge. The source of the artifacts lies in the effective treatment of the electrostatic interactions within and between the perturbed atoms and remaining (partial) charges in the simulated system. To treat the electrostatic interactions effectively, lattice-summation (LS) methods or cutoff schemes in combination with a reaction-field contribution are usually employed. Both methods render the charging component of the calculated free energies sensitive to essential parameters of the system like the cutoff radius or the box side lengths. Here, we discuss the results of three previously published studies of ligand binding. These studies presented estimates of binding free energies that were artifacted due to the charged nature of the ligands. We show that the size of the artifacts can be efficiently calculated and raw simulation data can be corrected. We compare the corrected results with experimental estimates and nonartifacted estimates from path-sampling methods. Although the employed correction scheme involves computationally demanding continuum-electrostatics calculations, we show that the correction estimate can be deduced from a small sample of configurations rather than from the entire ensemble. This observation makes the calculations of correction terms feasible for complex biological systems. To show the general applicability of the proposed procedure, we also present results where the correction scheme was used to correct independent free energies obtained from simulations employing a cutoff scheme or LS electrostatics. In this work, we give practical guidelines on how to apply the appropriate corrections easily.
Topics: Artifacts; Binding Sites; DNA; Distamycins; Ligands; Molecular Dynamics Simulation; Netropsin; Solvents; Static Electricity; Thermodynamics; Trypsin Inhibitors
PubMed: 31930547
DOI: 10.1002/jcc.26143