RESUMEN
7H-Dibenzo[c,g]carbazole (DBC) is a prevalent environmental contaminant that induces tumorigenesis in several experimental animals. Recently, it has been utilized to develop high-performance solar cells and organic phosphorescent materials. It is imperative to strengthen investigations of DBC metabolism to understand its potential risks to human health. In this study, human CYP1A1 was employed as the metabolic enzyme to investigate the metabolic mechanism of DBC by molecular docking, molecular dynamics (MD) simulation, and quantum mechanical (QM) calculation. The results indicate that DBC binds to CYP1A1 in two modes (mode 1 and mode 2) mainly through nonpolar solvation energies (ΔGnonpolar). The formation of the two binding modes is attributed to the anchoring effect of the hydrogen bond formed by DBC with Asp320 (mode 1) or Ser116 (mode 2). Mode 1 is a "reactive" conformation, while mode 2 is not considered a "reactive" conformation. C5 is identified as the dominant site, and the pyrrole nitrogen cannot participate in the metabolism. DBC is metabolized mainly by a distinct electrophilic addition-rearrangement mechanism, with an energy barrier of 21.74 kcal/mol. The results provide meaningful insights into the biometabolic process of DBC and contribute to understanding its environmental effects and health risks.
Asunto(s)
Carbazoles , Citocromo P-450 CYP1A1 , Simulación del Acoplamiento Molecular , Simulación de Dinámica Molecular , Carbazoles/química , Carbazoles/metabolismo , Citocromo P-450 CYP1A1/metabolismo , Citocromo P-450 CYP1A1/química , Humanos , Teoría Cuántica , Enlace de HidrógenoRESUMEN
Streptocarbazoles are a class of indolocarbazole (ICZ) compounds produced by Streptomyces strains that feature unique cyclic N-glycosidic linkages between the 1,3-carbon atoms of the glycosyl moiety and the two indole nitrogen atoms. Although several streptocarbazole compounds display effective cytotoxic activity, their biosynthesis remains unclear. Herein, through the inactivation of the aminotransferase gene spcI in the staurosporine biosynthetic gene cluster spc followed by heterologous expression, two new streptocarbazole derivatives (1 and 3) and three known ICZs (2, 4, and 5) were generated. Their structures were determined by a combination of spectroscopic methods, circular dichroism measurements, and single-crystal X-ray diï¬raction. Compounds 1-4 displayed moderate cytotoxicity against HCT-116 cell line, and compounds 3 and 4 were effective against Huh 7 cell line. Double-gene knockout experiments allowed us to propose a biosynthetic pathway for streptocarbazole productions. Furthermore, by overexpression of the involving key enzymes, the production of streptocarbazoles 1 and 3 were improved by approximately 1.5-2.5 fold. IMPORTANCE: Indolocarbazoles (ICZs) are a group of antitumor agents, with several analogs used in clinical trials. Therefore, the identification of novel ICZ compounds is important for drug discovery. Streptocarbazoles harbor unique N-glycosidic linkages (N13-C1' and N12-C3'), distinguishing them from the representative ICZ compound staurosporine; however, their biosynthesis remains unclear. In this study, two new streptocarbazoles (1 and 3) with cytotoxic activities were obtained by manipulating the staurosporine biosynthetic gene cluster spc followed by heterologous expression. The biosynthetic pathway of streptocarbazoles was proposed, and their productions were improved through the overexpression of the key enzymes involved. This study enriches the structural diversity of ICZ compounds and would facilitate the discovery of new streptocarbazoles via synthetic biological strategies.
Asunto(s)
Carbazoles , Streptomyces , Estaurosporina/farmacología , Carbazoles/farmacología , Carbazoles/química , Carbazoles/metabolismo , Streptomyces/metabolismo , Familia de MultigenesRESUMEN
To elucidate why plasmid-borne catabolic ability differs among host bacteria, we assessed the expression dynamics of the Pant promoter on the carbazole-degradative conjugative plasmid pCAR1 in Pseudomonas putida KT2440(pCAR1) (hereafter, KTPC) and Pseudomonas resinovorans CA10. The Pant promoter regulates the transcription of both the car and ant operons, which are responsible for converting carbazole into anthranilate and anthranilate into catechol, respectively. In the presence of anthranilate, transcription of the Pant promoter is induced by the AraC/XylS family regulator AntR, encoded on pCAR1. A reporter cassette containing the Pant promoter followed by gfp was inserted into the chromosomes of KTPC and CA10. After adding anthranilate, GFP expression in the population of CA10 showed an unimodal distribution, whereas a small population with low GFP fluorescence intensity appeared for KTPC. CA10 has a gene, antRCA, that encodes an iso-functional homolog of AntR on its chromosome. When antRCA was disrupted, a small population with low GFP fluorescence intensity appeared. In contrast, overexpression of pCAR1-encoded AntR in KTPC resulted in unimodal expression under the Pant promoter. These results suggest that the expression of pCAR1-encoded AntR is insufficient to ameliorate the stochastic expression of the Pant promoter. Raman spectra of single cells collected using deuterium-labeled carbazole showed that the C-D Raman signal exhibited greater variability for KTPC than CA10. These results indicate that heterogeneity at the transcriptional level of the Pant promoter due to insufficient AntR availability causes fluctuations in the pCAR1-borne carbazole-degrading capacity of host bacterial cells.IMPORTANCEHorizontally acquired genes increase the competitiveness of host bacteria under selective conditions, although unregulated expression of foreign genes may impose fitness costs. The "appropriate" host for a plasmid is empirically known to maximize the expression of plasmid-borne traits. In the case of pCAR1-harboring Pseudomonas strains, P. resinovorans CA10 exhibits strong carbazole-degrading capacity, whereas P. putida KT2440 harboring pCAR1 exhibits low degradation capacity. Our results suggest that a chromosomally encoded transcription factor affects transcriptional and metabolic fluctuations in host cells, resulting in different carbazole-degrading capacities as a population. This study may provide a clue for determining appropriate hosts for a plasmid and for regulating the expression of plasmid-borne traits, such as the degradation of xenobiotics and antibiotic resistance.
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Pseudomonas putida , Plásmidos/genética , Pseudomonas putida/genética , Pseudomonas putida/metabolismo , Regiones Promotoras Genéticas , Carbazoles/metabolismo , ortoaminobenzoatos/metabolismo , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismoRESUMEN
Cells of most eukaryotic species contain mitochondria, which play a role in physiological processes such as cellular senescence, metabolism, and autophagy. Viscosity is considered a key marker for many illnesses and is involved in several crucial physiological processes. Cyanide (CN-) can target cytochrome-c oxidase, disrupting the mitochondrial electron transport chain and causing cell death through asphyxiation. In this study, a fluorescent probe named HL-1, which targets mitochondria and measures viscosity and CN- levels, was designed and synthesized. HL-1 is viscosity-sensitive, with a linear correlation coefficient of up to 0.992. In addition, HL-1 was found to change color substantially during a nucleophilic addition reaction with CN-, which has a low detection limit of 47 nM. HL-1 not only detects viscosity and exogenous CN- in SKOV-3 cells and zebrafish but also monitors viscosity changes during mitochondrial autophagy in real time. Furthermore, HL-1 has been used successfully to monitor changes in mitochondrial membrane potential during apoptosis. Endogenous CN- in plant samples was quantified. HL-1 provides new ideas for studying viscosity and CN-.
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Colorantes Fluorescentes , Pez Cebra , Animales , Humanos , Colorantes Fluorescentes/metabolismo , Viscosidad , Cianuros , Mitocondrias/metabolismo , Células HeLa , Carbazoles/metabolismoRESUMEN
Soil ecosystems are being more contaminated with polyhalogenated carbazoles (PHCZs), which raising much attention about their impact on soil microorganisms. 3-Bromocarbazole (3-BCZ) and 1,3,6,8-tetrabromocarbazole (1,3,6,8-TBCZ) are two typical PHCZs with high detection rates in the soil environment. However, ecological risk research on these two PHCZs in soil is still lacking. In the present study, after 80 days of exposure, the ecological influence of 3-BCZ and 1,3,6,8-TBCZ was investigated based on 16S rDNA sequencing, ITS sequencing, gene (16S rDNA, ITS, amoA, nifH, narG and cbbL) abundance and soil enzyme activity. The results showed that the bacterial 16S rDNA gene abundance significantly decreased under 3-BCZ and 1,3,6,8-TBCZ exposure after 80 days of incubation. The fungal ITS gene abundance significantly decreased under 1,3,6,8-TBCZ (10 mg/kg) exposure. PHCZs contributed to the alteration of bacteria and fungi community abundance. Bacteria Sphingomonas, RB41 and fungus Mortierella, Cercophora were identified as the most dominant genera. The two PHCZs consistently decreased the relative abundance of Sphingomonas, Lysobacter, Dokdonella, Mortierella and Cercophora etc at 80th day. These keystone taxa are related to the degradation of organic compounds, carbon metabolism, and nitrogen metabolism and may thus have influence on soil ecological functions. Bacterial and fungal functions were estimated using functional annotation of prokaryotic taxa (FAPROTAX) and fungi functional guild (FUNGuild), respectively. The nitrogen and carbon metabolism pathway were affected by 3-BCZ and 1,3,6,8-TBCZ. The soil nitrogen-related functions of aerobic ammonia oxidation were decreased but the soil carbon-related functions of methanol oxidation, fermentation, and hydrocarbon degradation were increased at 80th day. The effects of 3-BCZ and 1,3,6,8-TBCZ on the abundances of the amoA, nifH, narG, and cbbL genes showed a negative trend. These results elucidate the ecological effects of PHCZs and extend our knowledge on the structure and function of soil microorganisms in PHCZ-contaminated ecosystems.
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Microbiota , Suelo , Carbazoles/metabolismo , Bacterias/genética , Bacterias/metabolismo , Nitrógeno , Carbono , ADN Ribosómico , Microbiología del SueloRESUMEN
Natural products such as indolocarbazoles are a valuable source of highly bioactive compounds with numerous potential applications in the pharmaceutical industry. Arcyriaflavin A, isolated from marine invertebrates and slime molds, is one representative of this group and acts as a cyclin D1-cyclin-dependent kinase 4 inhibitor. To date, access to this compound has mostly relied on multi-step total synthesis. In this study, biosynthetic access to arcyriaflavin A was explored using recombinant Pseudomonas putida KT2440 based on a previously generated producer strain. We used a Design of Experiment approach to analyze four key parameters, which led to the optimization of the bioprocess. By engineering the formation of outer membrane vesicles and using an adsorbent in the culture broth, we succeeded to increase the yield of arcyriaflavin A in the cell-free supernatant, resulting in a nearly eight-fold increase in the overall production titers. Finally, we managed to scale up the bioprocess leading to a final yield of 4.7â mg arcyriaflavin A product isolated from 1â L of bacterial culture. Thus, this study showcases an integrative approach to improve biotransformation and moreover also provides starting points for further optimization of indolocarbazole production in P. putida.
Asunto(s)
Pseudomonas putida , Pseudomonas putida/metabolismo , Triptófano/metabolismo , Carbazoles/metabolismo , BiotransformaciónRESUMEN
BACKGROUND: Storage of samples may be necessary prior to testing drug levels in certain study designs; however, the effect of storage duration on measured drug levels is not known for all drugs. OBJECTIVES: The objective of this study was to evaluate the stability of carprofen in canine plasma when stored at -80°C for 6 months. METHODS: Six healthy dogs were enrolled (1-10 years old, 17-35 kg) and received compounded carprofen at 2.2 mg/kg orally every 12 h for 2 days. On the third day, blood was collected immediately before the morning dose (trough), then 1 and 6 h after the dose (sampling timepoint). Whole blood was immediately centrifuged, and plasma was stored at -80°C. Plasma carprofen concentration was measured at day 2, week 2 and then monthly for 6 months using reversed-phase high-performance liquid chromatography. The measured carprofen concentrations were analysed statistically using a linear mixed effects model. RESULTS: There was no effect of storage time over 6 months (p = 0.891) on measured carprofen levels. Although there was an effect of sampling timepoint (0, 1 and 6 h) (p < 0.001), the interaction between storage timepoint and sampling timepoint was not statistically significant (p = 1). CONCLUSIONS: Carprofen-laden canine plasma samples can be stored for up to 6 months before analysis with no degradation in carprofen concentrations expected.
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Antiinflamatorios no Esteroideos , Carbazoles , Perros , Animales , Carbazoles/metabolismoRESUMEN
Aquatic ecosystems are being more contaminated with polyhalogenated carbazoles (PHCZs), which raising concerns about their impact on aquatic organisms. Lycopene (LYC) exhibits several beneficial properties for fish via enhance antioxidant defenses and improve immunity. In this study, we attempted to investigate the hepatotoxic effects of typical PHCZs 3, 6-dichlorocarbazole (3,6-DCCZ) and the protective mechanisms of LYC. In this study, we found that yellow catfish (Pelteobagrus fulvidraco) exposure to 3,6-DCCZ (1.2 mg/L) resulted in hepatic inflammatory infiltration and disordered hepatocyte arrangement. Besides, we observed that 3,6-DCCZ exposure resulted in hepatic reactive oxygen species (ROS) overproduction and excessive autophagosome accumulation, accompanied with inhibition of phosphatidylinositol-3-kinase (PI3K)/protein kinase B (AKT) pathway. Subsequently, we confirmed that 3,6-DCCZ exposure triggered hepatic uncontrolled inflammatory response via activation of nuclear factor-κB (NF-κB) pathway, along with decreased plasma complement C3 (C3) and complement C4 (C4) levels. Meanwhile, yellow catfish exposed to 3,6-DCCZ exhibit an increased hepatic apoptosis phenomenon, as evidenced by the elevated number of positive TUNEL cells and upregulated expression of caspase3 and cytochrome C (CytC). In contrast, LYC treatment could alleviate the 3,6-DCCZ-induced pathological changes, hepatic ROS accumulation, autophagy, inflammatory response and apoptosis. To sum up, this study provided the demonstration that LYC exerts hepatoprotective effects to alleviate 3,6-DCCZ-induced liver damage by inihibiting ROS/PI3K-AKT/NF-κB signaling in yellow catfish.
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Bagres , FN-kappa B , Animales , FN-kappa B/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Licopeno/farmacología , Fosfatidilinositol 3-Quinasas/metabolismo , Bagres/metabolismo , Carbazoles/metabolismo , Carbazoles/farmacología , Ecosistema , Hígado/metabolismoRESUMEN
The aryl hydrocarbon receptor (AhR) is a decisive regulatory ligand-dependent transcription factor. It binds highly diverse ligands, which can be categorized as either endogenous or exogenous. Ligand binding activates AhR, which can adjust inflammatory responses by modulating immune cells such as dendritic cells (DCs). However, how different AhR ligand classes impact the phenotype and function of human monocyte-derived DCs (hMoDCs) has not been extensively studied in a comparative manner. We, therefore, tested the effect of the representative compounds Benzo(a)pyrene (BP), 6-formylindolo[3,2-b]carbazole (FICZ), and Indoxyl 3-sulfate (I3S) on DC biology. Thereby, we reveal that BP significantly induces a tolerogenic response in lipopolysaccharide-matured DCs, which is not apparent to the same extent when using FICZ or I3S. While all three ligand classes activate AhR-dependent pathways, BP especially induces the expression of negative immune regulators, and subsequently strongly subverts the T cell stimulatory capacity of DCs. Using the CRISPR/Cas9 strategy we also prove that the regulatory effect of BP is strictly AhR-dependent. These findings imply that AhR ligands contribute differently to DC responses and incite further studies to uncover the mechanisms and molecules which are involved in the induction of different phenotypes and functions in DCs upon AhR activation.
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Regulación de la Expresión Génica , Receptores de Hidrocarburo de Aril , Humanos , Receptores de Hidrocarburo de Aril/genética , Receptores de Hidrocarburo de Aril/metabolismo , Ligandos , Carbazoles/farmacología , Carbazoles/metabolismo , Indicán/metabolismo , Células Dendríticas , BiologíaRESUMEN
Neuroinflammation is a leading cause for neurological disorders. Carbazole alkaloids, isolated from the medicinal plants of Murraya species (Rutaceae), have exhibited wide pharmacological activities particularly for neuroinflammation. However, its underlying cellular targets and molecular mechanisms still remain unclear. In current study, we found that murrayafoline A (MA), a carbazole alkaloid obtained from Murraya tetramera, potently inhibited the production of neuroinflammation mediators, such as nitric oxide (NO), TNF-α, IL-6 and IL-1ß in LPS-induced BV-2 microglial cells. Then, we performed thermal proteome profiling (TPP) strategy to identify Specificity protein 1 (Sp1) as a potential cellular target of MA. Moreover, we performed surface plasmon resonance (SPR), cellular thermal shift assay (CETSA) and drug affinity responsive target stability (DRATS) assays to confirm the direct interaction between MA and Sp1. Furthermore, we downregulated Sp1 expression in BV2 cells using siRNA transfection, and observed that Sp1 knockdown significantly antagonized MA-mediated inhibition of neuroinflammation mediator production. Meanwhile, Sp1 knockdown also markedly reversed MA-mediated inactivation of IKKß/NF-κB and p38/JNK MAPKs pathways. Finally, in vivo studies revealed that MA significantly suppressed the expression of Iba-1, TNF-α, and IL-6, while increased the number of Nissl bodies in the brains of LPS-induced mice. Taken together, our study demonstrated that MA exerted obvious anti-neuroinflammation effect by directly targeting Sp1, thereby inhibiting NF-κB and MAPK signaling pathways. Our findings also provided a promising direction of pharmacological targeting Sp1 for anti-neuroinflammation therapeutics as well as novel agent development.
Asunto(s)
Alcaloides , Antiinflamatorios , Carbazoles , Murraya , Enfermedades Neuroinflamatorias , Factor de Transcripción Sp1 , Animales , Ratones , Alcaloides/farmacología , Alcaloides/uso terapéutico , Antiinflamatorios/farmacología , Antiinflamatorios/uso terapéutico , Carbazoles/metabolismo , Carbazoles/uso terapéutico , Interleucina-6/metabolismo , Lipopolisacáridos , Microglía/efectos de los fármacos , Murraya/química , FN-kappa B/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Factor de Transcripción Sp1/metabolismo , Enfermedades Neuroinflamatorias/tratamiento farmacológicoRESUMEN
Carbazole and its derivatives are polycyclic aromatic heterocycles with unusual toxicity and mutagenicity. However, disposal of these polycyclic aromatic heterocycles remains a significant challenge. This study focused on efficient resource recovery from carbazole using an obligate aerobe, Sphingobium yanoikuyae XLDN2-5, in microbial fuel cells (MFCs). S. yanoikuyae XLDN2-5 successfully achieved carbazole degradation and simultaneously electricity generation in MFCs with a maximum power density of 496.8 mW m-2 and carbazole degradation rate of 100%. It is the first time that S. yanoikuyae XLDN2-5 was discovered as an electrochemically active bacterium with high extracellular electron transfer (EET) capability. Redox mediator analysis indicated that no self-produced redox mediators were found for S. yanoikuyae XLDN2-5 under analysis conditions, and the exogenous redox mediators used in this study did not promote its EET. The nanowires produced by S. yanoikuyae XLDN2-5 cells were found in the biofilm by morphology characterization and the growth process of the nanowires was consistent with the discharge process of the MFC. Conductivity determination further verified that the nanowires produced by S. yanoikuyae XLDN2-5 cells were electrically conductive. Based on these results, it is speculated that S. yanoikuyae XLDN2-5 may mainly utilize conductive nanowires produced by itself rather than redox mediators to meet the requirements of normal energy metabolism when it grows in the low dissolved oxygen zone of the anodic biofilm. These novel findings on the EET mechanism of S. yanoikuyae XLDN2-5 lay a foundation for further exploration of polycyclic aromatic heterocyclic pollutants treatment in electrochemical devices, which may create new biotechnology processes for these pollutants control.
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Fuentes de Energía Bioeléctrica , Contaminantes Ambientales , Sphingomonadaceae , Fuentes de Energía Bioeléctrica/microbiología , Carbazoles/metabolismo , Electricidad , Electrodos , Contaminantes Ambientales/metabolismo , Oxígeno/metabolismo , Sphingomonadaceae/metabolismoRESUMEN
Thiabendazole (TBZ), is a persistent fungicide/anthelminthic and a serious environmental threat. We previously enriched a TBZ-degrading bacterial consortium and provided first evidence for a Sphingomonas involvement in TBZ transformation. Here, using a multi-omic approach combined with DNA-stable isotope probing (SIP) we verified the key degrading role of Sphingomonas and identify potential microbial interactions governing consortium functioning. SIP and amplicon sequencing analysis of the heavy and light DNA fraction of cultures grown on 13 C-labelled versus 12 C-TBZ showed that 66% of the 13 C-labelled TBZ was assimilated by Sphingomonas. Metagenomic analysis retrieved 18 metagenome-assembled genomes with the dominant belonging to Sphingomonas, Sinobacteriaceae, Bradyrhizobium, Filimonas and Hydrogenophaga. Meta-transcriptomics/-proteomics and non-target mass spectrometry suggested TBZ transformation by Sphingomonas via initial cleavage by a carbazole dioxygenase (car) to thiazole-4-carboxamidine (terminal compound) and catechol or a cleaved benzyl ring derivative, further transformed through an ortho-cleavage (cat) pathway. Microbial co-occurrence and gene expression networks suggested strong interactions between Sphingomonas and a Hydrogenophaga. The latter activated its cobalamin biosynthetic pathway and Sphingomonas its cobalamin salvage pathway to satisfy its B12 auxotrophy. Our findings indicate microbial interactions aligning with the 'black queen hypothesis' where Sphingomonas (detoxifier, B12 recipient) and Hydrogenophaga (B12 producer, enjoying detoxification) act as both helpers and beneficiaries.
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Dioxigenasas , Fungicidas Industriales , Sphingomonas , Sphingomonas/genética , Sphingomonas/metabolismo , Tiabendazol/metabolismo , Fungicidas Industriales/metabolismo , Dioxigenasas/metabolismo , Biodegradación Ambiental , Bacterias/genética , Bacterias/metabolismo , Carbazoles/metabolismo , Vitamina B 12/metabolismoRESUMEN
Chronic hyperglycemia favours the formation of advanced glycation end products (AGEs) which are responsible of many diabetic vascular complications. Keeping in view the medicinal properties of the1,2,3-triazole-conjugated analogs, the present study was designed to evaluate the possible effect of carbazole-linked 1,2,3-triazoles 2-16 against glucose- and methylglyoxal-AGEs-induced inflammation in human THP-1 monocytes. In vitro antiglycation, and metabolic assays were used to determine antiglycation, and cytotoxicity activities. DCFH-DA, immunostaining, immunoblotting, and ELISA techniques were employed to study the ROS and levels of proinflammatory mediators in THP-1 monocytes. Among all the synthesized carbazole-linked 1,2,3 triazoles, compounds 2, 7, 8, and 11-16 showed antiglycation activity in glucose- and MGO-modified bovine serum albumin models, whereas parent compound 1 only exhibited activity in glucose-BSA model. The metabolic assay demonstrated the non-toxic profile of compounds 1-2, 11-13, and 15 up to 100 µM concentration in both HepG2 and THP-1 cell lines. We found that compounds 11-13, and 15 attenuated AGEs-induced ROS formation (P < 0.001), and halted NF-ĸB translocation (P < 0.001), likewise standard drugs, PDTC, rutin, and quercetin, in THP-1 monocytes. Among the derivatives, compounds 12, and 13 also suppressed the AGEs-induced elevation of COX-2 (P < 0.001) and PGE2 (P < 0.001). Our data show that the carbazole-linked triazoles 12, and 13 hampering the formation of glycation products, prevent the activation of AGEs-ROS-NF-κB signaling pathway, and limit the proinflammatory COX-2 protein, and PGE2 induction in human THP-1 monocytes. Both these compounds can thus serve as leads for further studies towards the treatment and prevention of diabetic vascular complications.
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Angiopatías Diabéticas , Piruvaldehído , Carbazoles/metabolismo , Carbazoles/farmacología , Ciclooxigenasa 2/metabolismo , Angiopatías Diabéticas/metabolismo , Dinoprostona/metabolismo , Glucosa/metabolismo , Productos Finales de Glicación Avanzada , Humanos , Monocitos/metabolismo , FN-kappa B/metabolismo , Piruvaldehído/farmacología , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal , Triazoles/farmacologíaRESUMEN
Halogenated carbazoles are a new class of persistent organic pollutants with dioxin-like toxicity, and this study focused on the microbial degradation of 1,3,6,8-tetrabromocarbazole. In this study, a novel 1,3,6,8-tetrabromocarbazole (1,3,6,8-TBCZ) degrading strain TB-1 was isolated from contaminated soil and identified as Achromobacter sp. based on its 16S rRNA gene sequence analysis, morphological, physiological, and biochemical characteristics. The soil sample was collected from a pharmaceutical factory in Suzhou, China. The strain was able to effectively degrade 1 mg L-1 1,3,6,8-TBCZ in 7 d at pH 7.0 and 30 °C with 80% degradation rate. During the process, the intermediate metabolites were identified as Tribromocarbazole, dibromocarbazole and bromocarbazole via gas chromatography mass spectrometry (GC-MS). The results indicated that strain TB-1 may contribute to the bioremediation of polyhalogenated carbazoles (PHCs) in contaminated environment.
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Achromobacter , Contaminantes del Suelo , Achromobacter/metabolismo , Biodegradación Ambiental , Carbazoles/análisis , Carbazoles/metabolismo , ARN Ribosómico 16S/genética , Suelo , Microbiología del Suelo , Contaminantes del Suelo/análisisRESUMEN
In patients with chronic kidney disease (CKD) and in animal models of CKD, the transcription factor Aryl Hydrocabon Receptor (AhR) is overactivated. In addition to the canonical AhR targets constituting the AhR signature, numerous other genes are regulated by this factor. We identified neuronal pentraxin 1 (NPTX1) as a new AhR target. Belonging to the inflammatory protein family, NPTX1 seems of prime interest regarding the inflammatory state observed in CKD. Endothelial cells were exposed to tryptophan-derived toxins, indoxyl sulfate (IS) and indole-3-acetic acid (IAA). The adenine mouse model of CKD was used to analyze NPTX1 expression in the burden of uremia. NPTX1 expression was quantified by RT-PCR and western blot. AhR involvement was analyzed using silencing RNA. We found that IS and IAA upregulated NPTX1 expression in an AhR-dependent way. Furthermore, this effect was not restricted to uremic indolic toxins since the dioxin 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) and the tryptophan photoproduct 6-formylindolo[3,2-b]carbazole (FICZ) do the same. In CKD mice, NPTX1 expression was increased in the aorta. Therefore, NPTX1 is a new target of AhR and further work is necessary to elucidate its exact role during CKD.
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Proteína C-Reactiva/metabolismo , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Triptófano/metabolismo , Animales , Carbazoles/metabolismo , Células Cultivadas , Modelos Animales de Enfermedad , Humanos , Indicán/metabolismo , Ácidos Indolacéticos/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Dibenzodioxinas Policloradas/metabolismo , Receptores de Hidrocarburo de Aril/metabolismo , Insuficiencia Renal Crónica/metabolismo , Toxinas Biológicas/metabolismo , Uremia/metabolismoAsunto(s)
Quinasa de Linfoma Anaplásico/análisis , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Proteínas de Unión a Calmodulina/análisis , Carbazoles/farmacología , Proteínas de la Membrana/análisis , Proteínas del Tejido Nervioso/análisis , Piperidinas/farmacología , Quinasa de Linfoma Anaplásico/sangre , Proteínas de Unión a Calmodulina/sangre , Carbazoles/metabolismo , Carbazoles/uso terapéutico , Femenino , Humanos , Proteínas de la Membrana/sangre , Persona de Mediana Edad , Proteínas del Tejido Nervioso/sangre , Piperidinas/metabolismo , Piperidinas/uso terapéutico , Inhibidores de Proteínas Quinasas/metabolismo , Inhibidores de Proteínas Quinasas/farmacología , Inhibidores de Proteínas Quinasas/uso terapéuticoRESUMEN
The actinomycete Lentzea aerocolonigenes produces the antitumor antibiotic rebeccamycin. In previous studies the rebeccamycin production was significantly increased by the addition of glass beads during cultivation in different diameters between 0.5 and 2 mm and the induced mechanical stress by the glass beads was proposed to be responsible for the increased production. Thus, this study was conducted to be a systematic investigation of different parameters for macroparticle addition, such as bead diameter, concentration, and density (glass and ceramic) as well as shaking frequency, for a better understanding of the particle-induced stress on L. aerocolonigenes. The induced stress for optimal rebeccamycin production can be estimated by a combination of stress energy and stress frequency. In addition, the macroparticle-enhanced cultivation of L. aerocolonigenes was combined with soy lecithin addition to further increase the rebeccamycin concentration. With 100 g L-1 glass beads in a diameter of 969 µm and 5 g L-1 soy lecithin a concentration of 388 mg L-1 rebeccamycin was reached after 10 days of cultivation, which corresponds to the highest rebeccamycin concentrations achieved in shake flask cultivations of L. aerocolonigenes stated in literature so far.
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Actinobacteria/crecimiento & desarrollo , Carbazoles/metabolismo , Vidrio , Lecitinas/farmacología , Estrés Mecánico , Lecitinas/metabolismoRESUMEN
Sirtuins are signaling hubs orchestrating the cellular response to various stressors with roles in all major civilization diseases. Sirtuins remove acyl groups from lysine residues of proteins, thereby controlling their activity, turnover, and localization. The seven human sirtuins, SirT1-7, are closely related in structure, hindering the development of specific inhibitors. Screening 170,000 compounds, we identify and optimize SirT1-specific benzoxazine inhibitors, Sosbo, which rival the efficiency and surpass the selectivity of selisistat (EX527). The compounds inhibit the deacetylation of p53 in cultured cells, demonstrating their ability to permeate biological membranes. Kinetic analysis of inhibition and docking studies reveal that the inhibitors bind to a complex of SirT1 and nicotinamide adenine dinucleotide, similar to selisistat. These new SirT1 inhibitors are valuable alternatives to selisistat in biochemical and cell biological studies. Their greater selectivity may allow the development of better targeted drugs to combat SirT1 activity in diseases such as cancer, Huntington's chorea, or anorexia.
Asunto(s)
Benzoxazinas/química , Sirtuina 1/antagonistas & inhibidores , Acetilación/efectos de los fármacos , Amidas/química , Benzoxazinas/metabolismo , Benzoxazinas/farmacología , Sitios de Unión , Carbazoles/química , Carbazoles/metabolismo , Línea Celular Tumoral , Evaluación Preclínica de Medicamentos , Humanos , Concentración 50 Inhibidora , Cinética , Simulación del Acoplamiento Molecular , NAD/química , NAD/metabolismo , Isoformas de Proteínas/antagonistas & inhibidores , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Proteínas Recombinantes/biosíntesis , Proteínas Recombinantes/química , Proteínas Recombinantes/aislamiento & purificación , Sirtuina 1/genética , Sirtuina 1/metabolismo , Relación Estructura-Actividad , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
ATPases Associated with Diverse Cellular Activity (AAA ATPase) are essential enzymes found in all organisms. They are involved in various processes such as DNA replication, protein degradation, membrane fusion, microtubule serving, peroxisome biogenesis, signal transduction, and the regulation of gene expression. Due to the importance of AAA ATPases, several researchers identified and developed small-molecule inhibitors against these enzymes. We discuss six AAA ATPases that are potential drug targets and have well-developed inhibitors. We compare available structures that suggest significant differences of the ATP binding pockets among the AAA ATPases with or without ligand. The distances from ADP to the His20 in the His-Ser-His motif and the Arg finger (Arg353 or Arg378) in both RUVBL1/2 complex structures bound with or without ADP have significant differences, suggesting dramatically different interactions of the binding site with ADP. Taken together, the inhibitors of six well-studied AAA ATPases and their structural information suggest further development of specific AAA ATPase inhibitors due to difference in their structures. Future chemical biology coupled with proteomic approaches could be employed to develop variant specific, complex specific, and pathway specific inhibitors or activators for AAA ATPase proteins.
Asunto(s)
ATPasas Asociadas con Actividades Celulares Diversas/metabolismo , Bibliotecas de Moléculas Pequeñas/química , ATPasas Asociadas con Actividades Celulares Diversas/antagonistas & inhibidores , Sitios de Unión , Carbazoles/química , Carbazoles/metabolismo , Humanos , Simulación de Dinámica Molecular , Neoplasias/tratamiento farmacológico , Neoplasias/patología , Pirazoles/química , Pirazoles/metabolismo , Pirimidinas/química , Pirimidinas/metabolismo , Quinazolinas/química , Quinazolinas/metabolismo , Bibliotecas de Moléculas Pequeñas/metabolismo , Bibliotecas de Moléculas Pequeñas/uso terapéutico , Zearalenona/análogos & derivados , Zearalenona/química , Zearalenona/metabolismoRESUMEN
We reviewed haematological investigations for 43 patients treated at a single centre with alectinib, an inhibitor of anaplastic lymphoma kinase (ALK) which is considered standard first-line treatment for patients with ALK-rearranged advanced non-small cell lung cancer. Ninety-five percent of patients developed marked acanthocytosis, echinocytosis and/or spheroacanthocytosis, not observable with prior treatment with other ALK-inhibitors. Anaemia developed in 73% of patients (38% <100 g/L, 8% <80 g/L), though definite new haemolysis was present in only 11%. Eosin-5-maleimide binding was reduced in all assessed patients, and increased membrane cholesterol was identified in one patient assessed with lattice light sheet microscopy. We have identified a previously undescribed phenomenon whereby alectinib induces red cell membrane abnormalities in nearly all patients through an unclear, but likely ALK-independent, mechanism, resulting in mild anaemia without universal haemolysis.