Integrative transcriptome and whole-genome bisulfite sequencing analyses of a temperature-sensitive albino tea plant cultivar.

Green tea made from albino buds and leaves has a strong umami taste and aroma. The cultivar 'Zhonghuang 2' (ZH2, Camellia sinensis) is a natural mutant with young shoots that are yellow in spring and green or yellow-green in summer. However, the mechanism of leaf color change remains unclear. Here, we found that young shoots of ZH2 were yellow at low temperature (LT) and green at high temperature (HT), indicating that ZH2 is a temperature-sensitive cultivar. Transmission electron microscopy analysis showed that the grana in the chloroplasts of young shoots grown at LT were poorly stacked, which caused a lack of photoreactions and chlorophyll. RNA-seq results showed 1279 genes differentially expressed in the young shoots grown at LT compared with those at HT, including genes related to cytochrome synthesis, chloroplast development, photosynthesis, and DNA methylation. A whole-genome bisulfite sequencing assay revealed that the dynamics of DNA methylation levels in the CG, CHG, and CHH contexts decreased under LT, and the change was most obvious in the CHH context. Furthermore, 72 genes showed significant changes in both expression and DNA methylation levels, and most of them were related to cytochrome synthesis, chloroplast development, photosynthesis, transcription factors, and signaling pathways. These results demonstrate that DNA methylation is involved in the LT-regulated albino processes of ZH2. Changes in DNA methylation levels were associated with changes in gene expression levels, affecting the structure and function of chloroplasts, which may have a phenotypic impact on shoot and leaf color.

Vitamin D modulation of brain-gut-virome disorder caused by polystyrene nanoplastics exposure in zebrafish (Danio rerio).

BACKGROUND: Many studies have investigated how nanoplastics (NPs) exposure mediates nerve and intestinal toxicity through a dysregulated brain-gut axis interaction, but there are few studies aimed at alleviating those effects. To determine whether and how vitamin D can impact that toxicity, fish were supplemented with a vitamin D-low diet and vitamin D-high diet. RESULTS: Transmission electron microscopy (TEM) showed that polystyrene nanoplastics (PS-NPs) accumulated in zebrafish brain and intestine, resulting in brain blood-brain barrier basement membrane damage and the vacuolization of intestinal goblet cells and mitochondria. A high concentration of vitamin D reduced the accumulation of PS-NPs in zebrafish brain tissues by 20% and intestinal tissues by 58.8% and 52.2%, respectively, and alleviated the pathological damage induced by PS-NPs. Adequate vitamin D significantly increased the content of serotonin (5-HT) and reduced the anxiety-like behavior of zebrafish caused by PS-NPs exposure. Virus metagenome showed that PS-NPs exposure affected the composition and abundance of zebrafish intestinal viruses. Differentially expressed viruses in the vitamin D-low and vitamin D-high group affected the secretion of brain neurotransmitters in zebrafish. Virus AF191073 was negatively correlated with neurotransmitter 5-HT, whereas KT319643 was positively correlated with malondialdehyde (MDA) content and the expression of cytochrome 1a1 (cyp1a1) and cytochrome 1b1 (cyp1b1) in the intestine. This suggests that AF191073 and KT319643 may be key viruses that mediate the vitamin D reduction in neurotoxicity and immunotoxicity induced by PS-NPs. CONCLUSION: Vitamin D can alleviate neurotoxicity and immunotoxicity induced by PS-NPs exposure by directionally altering the gut virome. These findings highlight the potential of vitamin D to alleviate the brain-gut-virome disorder caused by PS-NPs exposure and suggest potential therapeutic strategies to reduce the risk of NPs toxicity in aquaculture, that is, adding adequate vitamin D to diet. Video Abstract.

Whole genome sequencing of Wilsonomyces carpophilus, an incitant of shot hole disease in stone fruits: insights into secreted proteins of a necrotrophic fungal repository.

BACKGROUND: Shot hole is one of the important fungal diseases in stone fruits viz., peach, plum, apricot and cherry caused by Wilsonomyces carpophilus and almond among nut crops. Fungicides significantly decrease the disease. Pathogenicity studies proved a wide host range of the pathogen infecting all stone fruits and almond among the nut crops, however, the mechanism underlying host-pathogen interaction is still unknown. Molecular detection of the pathogen using polymerase chain reaction (PCR) based simple sequence repeat (SSR) markers is also unknown due to the unavailability of the pathogen genome. METHODS AND RESULTS: We examined the morphology, pathology and genomics of the Wilsonomyces carpophilus. Whole genome sequencing of the W. carpophilus was carried out by Illumina HiSeq and PacBio high throughput sequencing plate-forms through hybrid assembly. Constant selection pressure alters the molecular mechanism of the pathogen causing disease. The studies revealed that the necrotrophs are more lethal with a complex pathogenicity mechanism and little-understood effector repositories. The different isolates of necrotrophic fungus W. carpophilus causing shot hole in stone fruits namely peach, plum, apricot and cherry, and almonds among the nut crops showed a significant variation in their morphology, however, the probability value (p = 0.29) suggests in-significant difference in the pathogenicity. Here, we reported draft genome of W. carpophilus of size 29.9 Mb (Accession number: PRJNA791904). A total of 10,901 protein-coding genes were predicted, including heterokaryon incompatibility genes, cytochrome-p450 genes, kinases, sugar transporters among others. We found 2851 simple sequence repeats (SSRs), tRNAs, rRNAs and pseudogenes in the genome. The most prominent proteins showing necrotrophic lifestyle of the pathogen were hydrolases, polysaccharide-degrading enzymes, esterolytic, lipolytic, and proteolytic enzymes accounted for 225 released proteins. Among the 223 fungal species, top-hit species distribution revealed the majority of hits against the Pyrenochaeta species followed by Ascochyta rabiei and Alternaria alternata. CONCLUSION: Draft genome of W. carpophilus is 29.9 Mb based on Illumina HiSeq and PacBio hybrid assembly. The necrotrophs are more lethal with a complex pathogenicity mechanism. A significant variation in morphology was observed in different pathogen isolates. A total of 10,901 protein-coding genes were predicted in the pathogen genome including heterokaryon incompatibility, cytochrome-p450 genes, kinases and sugar transporters. We found 2851 SSRs, tRNAs, rRNAs and pseudogenes, and prominent proteins showing necrotrophic lifestyle such as hydrolases, polysaccharide-degrading enzymes, esterolytic, lipolytic and proteolytic enzymes. The top-hit species distribution were against the Pyrenochaeta spp. followed by Ascochyta rabiei.

The effect of not-anaerobicization and discolored bacteria on uranium reduction by Shewanella sp. RCRI7.

Shewanella sp. RCRI7 is a native strain capable of reducing uranium in anaerobic conditions. In order to employ this bacterium for the bioremediation, the mutual effects of uranium and the bacteria are studied in two different approaches. The optimal settings for the bacterial proliferation capacity and uranium reduction without anaerobicization of the environment, as well as the related effects of bioremediation and bacterial color under uranium-reducing conditions, have been investigated in this study. Uranium reduction procedure was analyzed using XRD, spectrophotometry and ICP-AES. In addition, the uranium's effect on the population of the first-generation of the bacteria as well as the color and growth of the second-generation were investigated using neobar lam and CFU (Colony Forming Unit), respectively. Uranium toxicity reduced the population of non-anaerobicized bacteria more than the anaerobicized bacteria after one day of incubation, while the amount of uranium extracted by the bacteria was almost the same. In both situations, the bacteria were able to reduce uranium after two weeks of incubation. In addition to the cell counts, uranium toxicity disrupts the growth and development of healthy second-generation anaerobicized bacteria, as created creamy-colored colonies grow slower than red-colored colonies. Furthermore, due to malfunctioning cytochromes, unlike red bacteria, creamy-colored bacteria were unable to extract the optimum amount of uranium. This study reveals that reduced uranium can be produced in a deprived environment without anaerobicization. Creamy-colored Shewanella can remove soluble uranium, however the most effective bacteria have red cytochromes. These findings represent a big step forward in the industrialization of uranium bioremediation.

Developmentally regulated mitochondrial biogenesis and cell death competence in maize pollen.

BACKGROUND: Cytoplasmic male sterility (CMS) is a maternally inherited failure to produce functional pollen that most commonly results from expression of novel, chimeric mitochondrial genes. In Zea mays, cytoplasmic male sterility type S (CMS-S) is characterized by the collapse of immature, bi-cellular pollen. Molecular and cellular features of developing CMS-S and normal (N) cytoplasm pollen were compared to determine the role of mitochondria in these differing developmental fates. RESULTS: Terminal deoxynucleotidyl transferase dUTP nick end labeling revealed both chromatin and nuclear fragmentation in the collapsed CMS-S pollen, demonstrating a programmed cell death (PCD) event sharing morphological features with mitochondria-signaled apoptosis in animals. Maize plants expressing mitochondria-targeted green fluorescent protein (GFP) demonstrated dynamic changes in mitochondrial morphology and association with actin filaments through the course of N-cytoplasm pollen development, whereas mitochondrial targeting of GFP was lost and actin filaments were disorganized in developing CMS-S pollen. Immunoblotting revealed significant developmental regulation of mitochondrial biogenesis in both CMS-S and N mito-types. Nuclear and mitochondrial genome encoded components of the cytochrome respiratory pathway and ATP synthase were of low abundance at the microspore stage, but microspores accumulated abundant nuclear-encoded alternative oxidase (AOX). Cytochrome pathway and ATP synthase components accumulated whereas AOX levels declined during the maturation of N bi-cellular pollen. Increased abundance of cytochrome pathway components and declining AOX also characterized collapsed CMS-S pollen. The accumulation and robust RNA editing of mitochondrial transcripts implicated translational or post-translational control for the developmentally regulated accumulation of mitochondria-encoded proteins in both mito-types. CONCLUSIONS: CMS-S pollen collapse is a PCD event coincident with developmentally programmed mitochondrial events including the accumulation of mitochondrial respiratory proteins and declining protection against mitochondrial generation of reactive oxygen species.

New Halogenated Compounds from Halimeda macroloba Seaweed with Potential Inhibitory Activity against Malaria.

Malaria is one of the most important infectious diseases worldwide. The causative of the most severe forms of malaria, Plasmodium falciparum, has developed resistances against all the available antimalarial drugs. In the present study, the phytochemical investigation of the green seaweed Halimeda macroloba has afforded two new compounds 1-2, along with 4 known ones 3-6. The structures of the compounds had been confirmed using 1& 2D-NMR and HRESIMS analyses. Extensive machine-learning-supported virtual-screening suggested cytochrome-C enzyme as a potential target for compound 2. Docking, absolute-binding-free-energy (ΔGbinding) and molecular-dynamics-simulation (MDS) of compound 2 revealed the strong binding interaction of this compound with cytochrome-C. In vitro testing for crude extract and isolated compounds revealed the potential in vitro inhibitory activity of both extract and compound 2 against P. falciparum. The crude extract was able to inhibit the parasite growth with an IC50 value of 1.8 ± 0.35 µg/mL. Compound 2 also showed good inhibitory activity with an IC50 value of 3.2 ± 0.23 µg/mL. Meanwhile, compound 6 showed moderate inhibitory activity with an IC50 value of 19.3 ± 0.51 µg/mL. Accordingly, the scaffold of compound 2 can be considered as a good lead compound for the future development of new antimalarial agents.

Bioenergetics and Reactive Nitrogen Species in Bacteria.

The production of reactive nitrogen species (RNS) by the innate immune system is part of the host's defense against invading pathogenic bacteria. In this review, we summarize recent studies on the molecular basis of the effects of nitric oxide and peroxynitrite on microbial respiration and energy conservation. We discuss possible molecular mechanisms underlying RNS resistance in bacteria mediated by unique respiratory oxygen reductases, the mycobacterial bcc-aa3 supercomplex, and bd-type cytochromes. A complete picture of the impact of RNS on microbial bioenergetics is not yet available. However, this research area is developing very rapidly, and the knowledge gained should help us develop new methods of treating infectious diseases.

Coupling methanogenesis with iron reduction by acetotrophic Methanosarcina mazei zm-15.

Application of ferric iron is conventionally considered to inhibit methanogenesis in anoxic environments. Here we show that Methanosarcina mazei zm-15, a strain isolated from the natural wetland of Tibetan plateau, is capable of Fe(III) reduction, which significantly promotes its growth and methanogenesis. We grew Ms. mazei zm-15 in a medium containing acetate supplemented with Fe(III) in ferric citrate or ferrihydrite and to some cultures anthraquinone-2,6-disulfonate (AQDS) was applied as an electron shuttle. The reduction of Fe(III) species occurred immediately. Ferric citrate was more readily reduced than ferrihydrite. The X-ray diffraction spectra analysis showed the formation of magnetite from ferrihydrite and amorphous reduced products from ferrihydrite plus AQDS. The analysis of protein contents revealed that Fe(III) reduction contributed 36%-46% of the cell growth. The growth yield, estimated as protein increment per acetate consumed for Fe(III) reduction, increased by 20- to 30-fold compared with methanogenesis, which is in consistence with the difference in free energy available by Fe(III) reduction relative to methanogenesis. We propose that the outer-surface multiheme c-type cytochrome predicted from Ms. mazei zm-15 genome serves as the terminal reductase with the energy-converting hydrogenase and F420 H2 dehydrogenase involved in electron transport chain for Fe(III) reduction. The findings shed a light to better understand the ecophysiology of Methanosarcina in anaerobic environments.

Interaction of picolinamide fungicide primary metabolites UK-2A and CAS-649 with the cytochrome bc1 complex Qi site: mutation effects and modelling in Saccharomyces cerevisiae.

BACKGROUND: Fenpicoxamid and florylpicoxamid are picolinamide fungicides targeting the Qi site of the cytochrome bc1 complex, via their primary metabolites UK-2A and CAS-649, respectively. We explore binding interactions and resistance mechanisms for picolinamides, antimycin A and ilicicolin H in yeast by testing effects of cytochrome b amino acid changes on fungicide sensitivity and interpreting results using molecular docking. RESULTS: Effects of amino acid changes on sensitivity to UK-2A and CAS-649 were similar, with highest resistance associated with exchanges involving G37 and substitutions N31K and L198F. These changes, as well as K228M, also affected antimycin A, while ilicicolin H was affected by changes at G37 and L198, as well as Q22E. N31 substitution patterns suggest that a lysine at position 31 introduces an electrostatic interaction with neighbouring D229, causing disruption of a key salt-bridge interaction with picolinamides. Changes involving G37 and L198 imply resistance primarily through steric interference. G37 changes also showed differences between CAS-649 and UK-2A or antimycin A with respect to branched versus unbranched amino acids. N31K and substitution of G37 by large amino acids reduced growth rate substantially while L198 substitutions showed little effect on growth. CONCLUSION: Binding of UK-2A and CAS-649 at the Qi site involves similar interactions such that general cross-resistance between fenpicoxamid and florylpicoxamid is anticipated in target pathogens. Some resistance mutations reduced growth rate and could carry a fitness penalty in pathogens. However, certain changes involving G37 and L198 carry little or no growth penalty and may pose the greatest risk for resistance development in the field. © 2022 Society of Chemical Industry.

Hepatoprotective effect of Sophora moorcroftiana (Benth.) Benth.Ex baker seeds in vivo and in vitro.

The leguminosae of Sophora moorcroftiana (Benth.) Benth.ex Baker is a drought-resistant endemic Sophora shrub species from the Qinghai-Tibet Plateau, and its seeds have hepatoprotective effects. To study the effect of S. moorcroftiana seeds on liver injury and the molecular mechanism underlying the beneficial effects, liquid chromatography-mass spectrometry was used to detect the main active components in the ethanol extract of S. moorcroftiana seeds (SM). Male mice were divided into six groups (n = 8): normal control (NC), CCl4, SM (50, 100, 200 mg/kg), and dimethyl diphenyl bicarboxylate (150 mg/kg) groups. Mice were treated as indicated (once/day, orally) for 14 days, and CCl4 (2 mL/kg) was administered intraperitoneally. The serum and liver of mice were used for biochemical assays. To explore the underlying mechanism, HepG2 cells were treated with SM, stimulated with tert-butyl hydroperoxide (t-BHP, 50 µM), and analyzed by Western blotting. The major active compounds of SM were alkaloids including 22 compounds. Serum alanine transaminase (ALT), aspartate transaminase (AST), and alkaline phosphatase (ALP) decreased in the SM (200 mg/kg) group. SM can activate the expression of pregnane X receptor (PXR) and downstream molecules cytochrome P4503A11 enzyme (CYP3A11), UDP glucuronosyltransferase 1 family polypeptide A 1 (UGT1A1), and inhibit the multidrug resistance protein 2 (MRP2). In addition, SM improved cell viability in t-BHP-induced HepG2 cells (64% to 83%) and decreased the activation of the mitogen-activated protein kinase (MAPK) pathway. The main compounds in SM were alkaloids. SM showed hepatoprotective effects possibly mediated by the suppression of oxidative stress through the MAPK pathway.

Effects of Exposure to Low Concentrations of Oil on the Expression of Cytochrome P4501a and Routine Swimming Speed of Atlantic Haddock (Melanogrammus aeglefinus) Larvae In Situ.

Exposure to environmentally relevant concentrations of oil could impact survival of fish larvae in situ through subtle effects on larval behavior. During the larval period, Atlantic haddock (Melanogrammus aeglefinus) are transported toward nursery grounds by ocean currents and active swimming, which can modify their drift route. Haddock larvae are sensitive to dispersed oil; however, whether exposure to oil during development impacts the ability of haddock larvae to swim in situ is unknown. Here, we exposed Atlantic haddock embryos to 10 and 80 µg oil/L (0.1 and 0.8 µg ∑PAH/L) of crude oil for 8 days and used a novel approach to measure its effect on the larval swimming behavior in situ. We assessed the swimming behavior of 138 haddock larvae in situ, in the North Sea, using a transparent drifting chamber. Expression of cytochrome P4501a (cyp1a) was also measured. Exposure to 10 and 80 µg oil/L significantly reduced the average in situ routine swimming speed by 30-40% compared to the controls. Expression of cyp1a was significantly higher in both exposed groups. This study reports key information for improving oil spill risk assessment models and presents a novel approach to study sublethal effects of pollutants on fish larvae in situ.

Differential gene content and gene expression for bacterial evolution and speciation of Shewanella in terms of biosynthesis of heme and heme-requiring proteins.

BACKGROUND: Most species of Shewanella harbor two ferrochelatase paralogues for the biosynthesis of c-type cytochromes, which are crucial for their respiratory versatility. In our previous study of the Shewanella loihica PV-4 strain, we found that the disruption of hemH1 but not hemH2 resulted in a significant accumulation of extracellular protoporphyrin IX (PPIX), but it is different in Shewanella oneidensis MR-1. Hence, the function and transcriptional regulation of two ferrochelatase genes, hemH1 and hemH2, are investigated in S. oneidensis MR-1. RESULT: In the present study, deletion of either hemH1 or hemH2 in S. oneidensis MR-1 did not lead to overproduction of extracellular protoporphyrin IX (PPIX) as previously described in the hemH1 mutants of S. loihica PV-4. Moreover, supplement of exogenous hemins made it possible to generate the hemH1 and hemH2 double mutant in MR-1, but not in PV-4. Under aerobic condition, exogenous hemins were required for the growth of MR-1ΔhemH1ΔhemH2, which also overproduced extracellular PPIX. These results suggest that heme is essential for aerobic growth of Shewanella species and MR-1 could also uptake hemin for biosynthesis of essential cytochrome(s) and respiration. Besides, the exogenous hemin mediated CymA cytochrome maturation and the cellular KatB catalase activity. Both hemH paralogues were transcribed in wild-type MR-1, and the hemH2 transcription was remarkably up-regulated in MR-1ΔhemH1 mutant to compensate for the loss of hemH1. The periplasmic glutathione peroxidase gene pgpD, located in the same operon with hemH2, and a large gene cluster coding for iron, heme (hemin) uptake systems are absent in the PV-4 genome. CONCLUSION: Our results indicate that the genetic divergence in gene content and gene expression between these Shewanella species, accounting for the phenotypic difference described here, might be due to their speciation and adaptation to the specific habitats (iron-rich deep-sea vent versus iron-poor freshwater) in which they evolved and the generated mutants could potentially be utilized for commercial production of PPIX.

Repeated evolution of cytochrome P450-mediated spiroketal steroid biosynthesis in plants.

Diosgenin is a spiroketal steroidal natural product extracted from plants and used as the single most important precursor for the world steroid hormone industry. The sporadic occurrences of diosgenin in distantly related plants imply possible independent biosynthetic origins. The characteristic 5,6-spiroketal moiety in diosgenin is reminiscent of the spiroketal moiety present in anthelmintic avermectins isolated from actinomycete bacteria. How plants gained the ability to biosynthesize spiroketal natural products is unknown. Here, we report the diosgenin-biosynthetic pathways in himalayan paris (Paris polyphylla), a monocot medicinal plant with hemostatic and antibacterial properties, and fenugreek (Trigonella foenum-graecum), an eudicot culinary herb plant commonly used as a galactagogue. Both plants have independently recruited pairs of cytochromes P450 that catalyze oxidative 5,6-spiroketalization of cholesterol to produce diosgenin, with evolutionary progenitors traced to conserved phytohormone metabolism. This study paves the way for engineering the production of diosgenin and derived analogs in heterologous hosts.

Inhibition of Rat CYP1A2 and CYP2C11 by Honokiol, a Component of Traditional Chinese Medicine.

BACKGROUND AND OBJECTIVES: Honokiol, a major constituent isolated from Magnolia officinalis, is regarded as a phytochemical marker and bioactive substance present in many traditional Chinese medicines. However, the effect of honokiol on cytochrome P450 (CYP) has not been thoroughly investigated. The aim of this study was to investigate the effect of honokiol on CYP1A2 and CYP2C11 in vitro and in vivo. METHODS: The effect of honokiol on CYP1A2 and CYP2C11 was investigated with rat liver microsomes (RLMs) by measuring phenacetin and tolbutamide metabolism (probe drugs for CYP1A2 and CYP2C11, respectively), and then explored in vivo by measuring the effect of honokiol (2.5 and 5 mg/kg, intravenous injection) on the pharmacokinetics of theophylline and tolbutamide (probe drugs for CYP1A2 and CYP2C11, respectively) in rats in vivo. RESULTS: Honokiol inhibited the formation of acetaminophen from phenacetin and 4-hydroxytolbutamide from tolbutamide in RLMs, with inhibition constant (Ki) values of 1.6 µM and 16.5 µM, respectively. In vivo, honokiol (2.5 or 5.0 mg/kg) increased the half-life (t1/2) of theophylline by 40.9% and 119.9%, decreased the clearance (CL) by 23.8% and 42.9%, and increased the area under the curve (AUC) by 41.3% and 83.4%, respectively. Similarly, the t1/2 of tolbutamide increased by 25.5% and 33.8%, the CL decreased by 14.3% and 19.1%, and the AUC increased by 19.2% and 25.7%, respectively. CONCLUSION: The inhibition of CYP1A2 by honokiol is greater than the inhibition of CYP2C11. The changes in the pharmacokinetics of theophylline and tolbutamide in rats treated with honokiol are due to the inhibition of CYP1A2 and CYP2C11 activity in a dose-dependent manner.

Transcriptome Profiling of Two Ornamental and Medicinal Papaver Herbs.

The Papaver spp. (Papaver rhoeas (Corn poppy) and Papaver nudicaule (Iceland poppy)) genera are ornamental and medicinal plants that are used for the isolation of alkaloid drugs. In this study, we generated 700 Mb of transcriptome sequences with the PacBio platform. They were assembled into 120,926 contigs, and 1185 (82.2%) of the benchmarking universal single-copy orthologs (BUSCO) core genes were completely present in our assembled transcriptome. Furthermore, using 128 Gb of Illumina sequences, the transcript expression was assessed at three stages of Papaver plant development (30, 60, and 90 days), from which we identified 137 differentially expressed transcripts. Furthermore, three co-occurrence heat maps are generated from 51 different plant genomes along with the Papaver transcriptome, i.e., secondary metabolite biosynthesis, isoquinoline alkaloid biosynthesis (BIA) pathway, and cytochrome. Sixty-nine transcripts in the BIA pathway along with 22 different alkaloids (quantified with LC-QTOF-MS/MS) were mapped into the BIA KEGG map (map00950). Finally, we identified 39 full-length cytochrome transcripts and compared them with other genomes. Collectively, this transcriptome data, along with the expression and quantitative metabolite profiles, provides an initial recording of secondary metabolites and their expression related to Papaver plant development. Moreover, these profiles could help to further detail the functional characterization of the various secondary metabolite biosynthesis and Papaver plant development associated problems.

The Eponymous Cofactors in Cytochrome P460s from Ammonia-Oxidizing Bacteria Are Iron Porphyrinoids Whose Macrocycles Are Dibasic.

The enzymes hydroxylamine oxidoreductase and cytochrome (cyt) P460 contain related unconventional "heme P460" cofactors. These cofactors are unusual in their inclusion of nonstandard cross-links between amino acid side chains and the heme macrocycle. Mutagenesis studies performed on the Nitrosomonas europaea cyt P460 that remove its lysine-heme cross-link show that the cross-link is key to defining the spectroscopic properties and kinetic competence of the enzyme. However, exactly how this cross-link confers these features remains unclear. Here we report the 1.45 Å crystal structure of cyt P460 from Nitrosomonas sp. AL212 and conclude that the cross-link does not lead to a change in hybridization of the heme carbon participating in the cross-link but rather enforces structural distortions to the macrocycle away from planarity. Time-dependent density functional theory coupled to experimental structural and spectroscopic analysis suggest that this geometric distortion is sufficient to define the spectroscopic properties of the heme P460 cofactor and provide clues toward establishing a relationship between heme P460 electronic structure and function.

Aging, Genetic Variations, and Ethnopharmacology: Building Cultural Competence Through Awareness of Drug Responses in Ethnic Minority Elders.

Unique drug responses that may result in adverse events are among the ethnocultural differences described by the Agency for Healthcare Research and Quality. These differences, often attributed to a lack of adherence on the part of the older adult, may be linked to genetic variations that influence drug responses in different ethnic groups. The paucity of research coupled with a lack of knowledge among health care providers compound the problem, contributing to further disparities, especially in this era of personalized medicine and pharmacogenomics. This article examines how age-related changes and genetic differences influence variations in drug responses among older adults in unique ethnocultural groups. The article starts with an overview of age-related changes and ethnopharmacology, moves to describing genetic differences that affect drug responses, with a focus on medications commonly prescribed for older adults, and ends with application of these issues to culturally congruent health care.

Creation of a gold nanoparticle based electrochemical assay for the detection of inhibitors of bacterial cytochrome bd oxidases.

Cytochrome bd oxidases are membrane proteins expressed by bacteria including a number of pathogens, which make them an attractive target for the discovery of new antibiotics. An electrochemical assay is developed to study the activity of these proteins and inhibition by quinone binding site tool compounds. The setup relies on their immobilization at electrodes specifically modified with gold nanoparticles, which allows achieving a direct electron transfer to/from the heme cofactors of this large enzyme. After optimization of the protein coverages, the assay shows at pH7 a good reproducibility and readout stability over time, and it is thus suitable for further screening of small molecule collections.

Effects of notoginsenoside R1 on CYP1A2, CYP2C11, CYP2D1, and CYP3A1/2 activities in rats by cocktail probe drugs.

CONTEXT: Notoginsenoside R1 (NGR1) is the main component with cardiovascular activity in Panax notoginseng (Burk.) F. H. Chen, an herbal medicine that is widely used to enhance blood circulation and dissipate blood stasis. OBJECTIVE: The objective of this study is to investigate NGR1's effects on CYP1A2, CYP2C11, CYP2D1, and CYP3A1/2 activities in rats in vivo through the use of the Cytochrome P450 (CYP450) probe drugs. MATERIALS AND METHODS: After pretreatment with NGR1 or physiological saline, the rats were administered intraperitoneally with a mixture solution of cocktail probe drugs containing caffeine (10 mg/kg), tolbutamide (15 mg/kg), metoprolol (20 mg/kg), and dapsone (10 mg/kg). The bloods were then collected at a set of time-points for the ultra-performance liquid chromatography/tandem mass spectrometry (UPLC-MS/MS) analysis. RESULTS: NGR1 was shown to exhibit an inhibitory effect on CYP1A2 by increased caffeine Cmax (43.13%, p < 0.01) and AUC0 - ∞ (40.57%, p < 0.01), and decreased CL/F (62.16%, p < 0.01) in the NGR1-treated group compared with those of the control group, but no significant changes in pharmacokinetic parameters of tolbutamide, metoprolol, and dapsone were observed between the two groups, indicating that NGR1 had no effects on rat CYP2C11, CYP2D1, and CYP3A1/2. DISCUSSION AND CONCLUSION: When NGR1 is co-administered with drugs that are metabolized by CYP1A2, the pertinent potential herb-drug interactions should be monitored.

Ephedra water decoction and cough tablets containing ephedra and liquorice induce CYP1A2 but not CYP2E1 hepatic enzymes in rats.

1. Ephedra water decoction (EWD) and cough tablets containing ephedra and liquorice (maxing cough tablets, MXCT) have been widely used in the treatment of asthma. In the clinic, EWD and MXCT may be prescribed with theophylline, one of the most popular antiasthmatic drugs. CYP1A2 and CYP2E1 are mainly involved in the oxidative metabolism of theophylline in human liver. Drug interactions involving the cytochrome P450 (CYP) isoforms generally are of two types: enzyme induction or enzyme inhibition. Enzyme inhibition reduces metabolism, whereas induction can increase it. 2. To evaluate the pretreatment effect of EWD and MXCT on CYP1A2 and CYP2E1, CYP1A2 and CYP2E1 activity, the protein expression and mRNA expression levels were determined. After pretreatment with EWD or MXCT, the enzyme activity, mRNA expression and protein expression of CYP1A2 were increased significantly (p < 0.05), but enzyme activity of CYP2E1 did not change compared with the control. 3. It was demonstrated that EWD or MXCT pretreatment obviously induced CYP1A2, therefore, in patients taking EWD or MXCT, possible CYP-induced drug interaction should be noted to decrease the risk of therapeutic failure or adverse effects resulting from the use of additional therapeutic agents.