Metabolism of xenobiotic compounds by enzymes in cell extracts of the fungus Cunninghamella elegans.
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ABSTRACT:
SUBMITTER: Wackett LP
PROVIDER: S-EPMC1158453 | biostudies-other | 1982 Jul
REPOSITORIES: biostudies-other
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Project description:The fungus Cunninghamella elegans is recognised as a microbial model of mammalian drug metabolism owing to its ability to catabolise xenobiotic compounds in an analogous fashion to animals. Its ability to produce phase I (oxidative) metabolites of drugs is associated with cytochrome P450 (CYP) activity; however, almost nothing is known about these enzymes in the fungus. In this paper we report the in silico analysis of the genome sequence of C. elegans B9769, which contains 32 genes putatively coding for CYPs. Based on their predicted amino acid sequences these were classified as belonging to CYP509, 5203, 5208, 5313, 5210, 61 and 51 families. Reverse transcription-quantitative PCR revealed that the gene coding for CYP5313D1 was significantly upregulated when C. elegans DSM1908 was cultivated in sabouraud dextrose in contrast to its expression in cells grown in Roswell Park Memorial Institute medium. This corresponded to the fungus' xenobiotic biotransformation ability when grown in the two media. Heterologous expression of cyp5313D1 in Pichia pastoris resulted in a recombinant strain that biotransformed flurbiprofen to 4'-hydroxyflurbiprofen, the same metabolite generated by C. elegans cultures. This is the first report of a xenobiotic-biotransforming CYP from this biotechnologically important fungus.
| S-EPMC6592952 | biostudies-literature
Project description:We investigated the metabolic capabilities of C. elegans using compounds whose metabolism has been well characterised in mammalian systems. We find that similar metabolites are produced in C. elegans as in mammals but that C. elegans is deficient in CYP1-like metabolism, as has been seen in other studies. We show that CYP-34A9, CYP-34A10 and CYP-36A1 are the principal enzymes responsible for the metabolism of tolbutamide in C. elegans. These are related to the mammalian enzymes that metabolise this compound but are not the closest homologs suggesting that sequence comparison alone will not predict functional conservation among cytochrome P450s. In mammals, metabolite production from amytryptiline and dextromethorphan is dependent on specific cytochrome P450s. However, in C. elegans we did not find evidence of similar specificity: the same metabolites were produced but in small amounts by numerous cytochrome P450s. We conclude that, while some aspects of cytochrome P450 mediated metabolism in C. elegans are similar to mammals, there are differences in the production of some metabolites and in the underlying genetics of metabolism.
| S-EPMC6127299 | biostudies-literature
Project description:Trematode infections occur worldwide causing considerable deterioration of human health and placing a substantial financial burden on the livestock industry. The hundreds of millions of people afflicted with trematode infections rely entirely on only two drugs (praziquantel and triclabendazole) for treatment. An understanding of anthelmintic biotransformation pathways in parasites should clarify factors that can modulate therapeutic potency of anthelmintics currently in use and may lead to the discovery of synergistic compounds for combination treatments. Despite the pronounced epidemiological significance of trematodes, there is still no adequate understanding of the functionality of their metabolic systems, including xenobiotic-metabolizing enzymes. The review is focused on the structure and functional significance of the xenobiotic-metabolizing system in trematodes. Knowledge in this field can solve practical problems related to the search for new targets for antiparasitic therapy based on a focused action on certain elements of the parasite's metabolic system. Knowledge of the functionality of this system is required to understand the adaptation of the biochemical processes of parasites residing in the host and mechanisms of drug resistance development, as well as to select a promising molecular target for the discovery and development of new anthelmintic drugs.
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Project description:BackgroundThe liver plays a major role in the metabolic activation of xenobiotics (drugs, chemicals such as pollutants, pesticides, food additives...). Among environmental contaminants of concern, heterocyclic aromatic amines (HAA) are xenobiotics classified by IARC as possible or probable carcinogens (2A or 2B). There exist little information about the effect of these HAA in humans. While HAA is a family of more than thirty identified chemicals, the metabolic activation and possible DNA adduct formation have been fully characterized in human liver for only a few of them (MeIQx, PhIP, A[Formula: see text]C).ResultsWe have developed a modeling approach in order to predict all the possible metabolites of a xenobiotic and enzymatic profiles that are linked to the production of metabolites able to bind DNA. Our prediction of metabolites approach relies on the construction of an enriched and annotated map of metabolites from an input metabolite.The pipeline assembles reaction prediction tools (SyGMa), sites of metabolism prediction tools (Way2Drug, SOMP and Fame 3), a tool to estimate the ability of a xenobotics to form DNA adducts (XenoSite Reactivity V1), and a filtering procedure based on Bayesian framework. This prediction pipeline was evaluated using caffeine and then applied to HAA. The method was applied to determine enzymes profiles associated with the maximization of metabolites derived from each HAA which are able to bind to DNA. The classification of HAA according to enzymatic profiles was consistent with their chemical structures.ConclusionsOverall, a predictive toxicological model based on an in silico systems biology approach opens perspectives to estimate the genotoxicity of various chemical classes of environmental contaminants. Moreover, our approach based on enzymes profile determination opens the possibility of predicting various xenobiotics metabolites susceptible to bind to DNA in both normal and physiopathological situations.
| S-EPMC8454073 | biostudies-literature
Project description:The US EPA's ToxCast program is designed to assess chemical perturbations of molecular and cellular endpoints using a variety of high-throughput screening (HTS) assays. However, existing HTS assays have limited or no xenobiotic metabolism which could lead to false positive (chemical is detoxified in vivo) as well as false negative results (chemical is bioactivated in vivo) and thus potential mischaracterization of chemical hazard. To address this challenge, the ten most prevalent human liver cytochrome P450 (CYP) enzymes were introduced into a human cell line (HEK293T) with low endogenous metabolic capacity. The CYP enzymes were introduced via transfection of modified mRNAs as either singlets or as a mixture in relative proportions as expressed in human liver. Initial experiments using luminogenic substrates demonstrate that CYP enzyme activities are significantly increased when co-transfected with an mRNA encoding a CYP accessory protein, P450 oxidoreductase (POR). Transfected HEK293T cells demonstrate the ability to produce predicted metabolites following treatment with well-studied CYP substrates for at least 18 h post-treatment. As a demonstration of how this method can be used to retrofit existing HTS assays, a proof-of-concept screen for cytotoxicity in HEK293T cells was conducted using 56 test compounds. The results demonstrate that the xenobiotic metabolism conferred by transfection of CYP-encoding mRNAs shifts the dose-response relationship for some of the tested chemicals such as aflatoxin B1 (bioactivation) and fenazaquin (detoxification). Overall, transfection of CYP-encoding mRNAs is an effective and portable solution for retrofitting existing cell-based HTS assays with metabolic competence.
| S-EPMC8171331 | biostudies-literature
Project description:Endophytic fungi are now recognized as sources of pharmacologically beneficial, novel bioactive compounds. This study was carried out to evaluate antiproliferative and antioxidative potential of a seaweed endophytic fungus Talaromyces purpureogenus. Extracts with different solvents of the fungus grown on different liquid media were assayed for the antiproliferative and antioxidative activities. Tested 6 cancer cell lines, the highest antiproliferative activity was observed in ethyl acetate extract of total culture grown in Potato Dextrose Broth for 28 days in a dose-dependent manner. The highest antioxidative activity was observed in hexane extract of fungal culture grown in Malt Extract Broth for 21 days. Analyzed for secondary metabolites, the extract revealed the presence of phenolics, alkaloids, flavonoids, steroids and terpenoids. Further, Gas Chromatography Mass Spectroscopy (GCMS) analysis of the extract revealed the presence of several compounds including 3-nitropropanoic acid, 4H-pyran-4-one 5-hydroxy-2-(hydroxymethyl), hexadecanoic acid, and octadecanoic acid, known to be cytotoxic or antioxidative. Among different cell lines tested, HeLa cells were the most vulnerable to the treatment of the fungal extract with an IC50 value of 101 ± 1 ?g/mL. The extract showed no significant cytotoxicity to the normal human embryonic kidney cell line (HEK 293 T) in the MTT assay. The ethyl acetate extract induced membrane damage and mitochondrial depolarization and thereby apoptosis and cytotoxicity in HeLa cells. The study marks marine-derived endophytes as potential sources for discovery of novel drugs.
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Project description:With a yearly production of about 39 million tons, brewer's spent grain (BSG) is the most abundant brewing industry byproduct. Because it is rich in fiber and protein, it is commonly used as cattle feed but could also be used within the human diet. Additionally, it contains many bioactive substances such as hydroxycinnamic acids that are known to be antioxidants and potent inhibitors of enzymes of glucose metabolism. Therefore, our study aim was to prepare different extracts-A1-A7 (solid-liquid extraction with 60% acetone); HE1-HE6 (alkaline hydrolysis followed by ethyl acetate extraction) and HA1-HA3 (60% acetone extraction of alkaline residue)-from various BSGs which were characterized for their total phenolic (TPC) and total flavonoid (TFC) contents, before conducting in vitro studies on their effects on the glucose metabolism enzymes α-amylase, α-glucosidase, dipeptidyl peptidase IV (DPP IV), and glycogen phosphorylase α (GPα). Depending on the extraction procedures, TPCs ranged from 20-350 µg gallic acid equivalents/mg extract and TFCs were as high as 94 µg catechin equivalents/mg extract. Strong inhibition of glucose metabolism enzymes was also observed: the IC50 values for α-glucosidase inhibition ranged from 67.4 ± 8.1 µg/mL to 268.1 ± 29.4 µg/mL, for DPP IV inhibition they ranged from 290.6 ± 97.4 to 778.4 ± 95.5 µg/mL and for GPα enzyme inhibition from 12.6 ± 1.1 to 261 ± 6 µg/mL. However, the extracts did not strongly inhibit α-amylase. In general, the A extracts from solid-liquid extraction with 60% acetone showed stronger inhibitory potential towards a-glucosidase and GPα than other extracts whereby no correlation with TPC or TFC were observed. Additionally, DPP IV was mainly inhibited by HE extracts but the effect was not of biological relevance. Our results show that BSG is a potent source of α-glucosidase and GPα inhibitors, but further research is needed to identify these bioactive compounds within BSG extracts focusing on extracts from solid-liquid extraction with 60% acetone.
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Project description:Humans are exposed to numerous xenobiotics, a majority of which are in the form of pharmaceuticals. Apart from human enzymes, recent studies have indicated the role of the gut bacterial community (microbiome) in metabolizing xenobiotics. However, little is known about the contribution of the plethora of gut microbiome in xenobiotic metabolism. The present study reports the results of analyses on xenobiotic metabolizing enzymes in various human gut microbiomes. A total of 397 available gut metagenomes from individuals of varying age groups from 8 nationalities were analyzed. Based on the diversities and abundances of the xenobiotic metabolizing enzymes, various bacterial taxa were classified into three groups, namely, least versatile, intermediately versatile and highly versatile xenobiotic metabolizers. Most interestingly, specific relationships were observed between the overall drug consumption profile and the abundance and diversity of the xenobiotic metabolizing repertoire in various geographies. The obtained differential abundance patterns of xenobiotic metabolizing enzymes and bacterial genera harboring them, suggest their links to pharmacokinetic variations among individuals. Additional analyses of a few well studied classes of drug modifying enzymes (DMEs) also indicate geographic as well as age specific trends.
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Project description:Extractions of the underground parts of valerian were prepared with water and ethanol (25-95%) at 25-75 °C. Extraction yields, bioactive compounds, and the 1,1-Diphenyl-2-picrylhydrazyl (DPPH) radical scavenging ability of lyophilized extracts were determined. The inhibitory effects of the extracts, valerenic acid derivatives and phenolic acids, on metabolic syndrome (MS)-related enzymes activities were further examined. Both roots and rhizomes extracted with 95% ethanol at 75 °C had the highest levels of bioactive compounds. The antioxidant capacity and inhibition of MS-related enzymes of the roots extract were better than those of the rhizomes. The roots extract more strongly inhibited pancreatic lipase (inhibition of 50% of enzyme activity (IC50), 17.59 mg/mL), angiotensin-converting enzyme (ACE, IC50, 3.75 mg/mL), α-amylase (IC50, 12.53 mg/mL), and α-glucosidase (IC50, 15.40 mg/mL). These four phenolic acids inhibited the activity of MS-related enzymes. Valerenic acid demonstrated more of an inhibitory ability for ACE (IC50, 0.225 mg/mL, except for caffeic acid) and α-glucosidase (IC50, 0.617 mg/mL) than phenolic acids. Valerian extract inhibited key enzyme activities that were associated with obesity (lipase), hypertension (ACE), and type 2 diabetes (α-amylase and α-glucosidase), suggesting that it is a potential candidate for the development of functional supplements.
| S-EPMC9914926 | biostudies-literature
Project description:Xenobiotics are ubiquitous in the environment and modified in the human body by phase I and II metabolism. Liquid chromatography coupled to high resolution mass spectrometry is a powerful tool to investigate these biotransformation products. We present a workflow based on stable isotope-assisted metabolomics and the bioinformatics tool MetExtract II for deciphering xenobiotic metabolites produced by human cells. Its potential was demonstrated by the investigation of the metabolism of deoxynivalenol (DON), an abundant food contaminant, in a liver carcinoma cell line (HepG2) and a model for colon carcinoma (HT29). Detected known metabolites included DON-3-sulfate, DON-10-sulfonate 2, and DON-10-glutathione as well as DON-cysteine. Conjugation with amino acids and an antibiotic was confirmed for the first time. The approach allows the untargeted elucidation of human xenobiotic products in tissue culture. It may be applied to other fields of research including drug metabolism, personalized medicine, exposome research, and systems biology to better understand the relevance of in vitro experiments.
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