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Vitamin K epoxide reductases (VKORs) are a large family of integral membrane enzymes found from bacteria to humans. Human VKOR, specific target of warfarin, has both the epoxide and quinone reductase activity to maintain the vitamin K cycle. Bacterial VKOR homologs, however, are insensitive to warfarin inhibition and are quinone reductases incapable of epoxide reduction. What affords the epoxide reductase activity in human VKOR remains unknown. Here, we show that a representative bacterial VKOR homolog can be converted to an epoxide reductase that is also inhibitable by warfarin. To generate this new activity, we first substituted several regions surrounding the active site of bacterial VKOR by those from human VKOR based on comparison of their crystal structures. Subsequent systematic substitutions narrowed down to merely eight residues, with the addition of a membrane anchor domain, that are responsible for the epoxide reductase activity. Substitutions corresponding to N80 and Y139 in human VKOR provide strong hydrogen bonding interactions to facilitate the epoxide reduction. The rest of six substitutions increase the size and change the shape of the substrate‐binding pocket, and the membrane anchor domain stabilizes this pocket while allowing certain flexibility for optimal binding of the epoxide substrate. Overall, our study reveals the structural features of the epoxide reductase activity carried out by a subset of VKOR family in the membrane environment. [ABSTRACT FROM AUTHOR]

Background: The mammalian major histocompatibility complex (MHC) harbours clusters of genes associated with the immunological defence of animals against infectious pathogens. At present, no complete MHC physical map is available for any of the wild ruminant species in the world. Results: The high-density physical map is composed of two contigs of 47 overlapping bacterial artificial chromosome (BAC) clones, with an average of 115 Kb for each BAC, covering the entire addax MHC genome. The first contig has 40 overlapping BAC clones covering an approximately 2.9 Mb region of MHC class I, class III, and class IIa, and the second contig has 7 BAC clones covering an approximately 500 Kb genomic region that harbours MHC class IIb. The relative position of each BAC corresponding to the MHC sequence was determined by comparative mapping using PCR screening of the BAC library of 192,000 clones, and the order of BACs was determined by DNA fingerprinting. The overlaps of neighboring BACs were cross-verified by both BAC-end sequencing and co-amplification of identical PCR fragments within the overlapped region, with their identities further confirmed by DNA sequencing. Conclusions: We report here the successful construction of a high-quality physical map for the addax MHC region using BACs and comparative mapping. The addax MHC physical map we constructed showed one gap of approximately 18 Mb formed by an ancient autosomal inversion that divided the MHC class II into IIa and IIb. The autosomal inversion provides compelling evidence that the MHC organizations in all of the ruminant species are relatively conserved. [ABSTRACT FROM AUTHOR]

Highlights • CRISPR/Cas9 induces random deletions in the target region of GABRR2 gene. • Both big and small indels can lead to unexpected high probability of exon truncation/skipping and exon recruitment. • It is the first observation of exon recruitment by CRISPR/Cas9-mediated GABRR2 gene editing. Abstract CRISPR/Cas9 and its variations provide an efficient tool for targeted genome editing. CRISPR/Cas9-based genome editing can induce mutations in genome that has high probability to cause exon truncation or deletion. However, screening mutations in diploid cells is difficult because of two copies of chromosome. It is an important task for determining genotypes in diploid cells subjected to editing before using these cells in gene function study. In this study, we applied CRISPR/Cas9 to edit the GABRR2 gene in mouse retinal ganglion cells to study what exactly happened in two alleles and what real mRNA isoforms formed in diploid cells. A single sgRNA was employed to generate double-strand DNA breaks. PCR sequencing was used for single clone validation in diploid cells subjected to editing. The indels and the corresponding effects at the target locus were further studied at genomic and RNA levels. We observed that CRISPR/Cas9 induces random deletions in the target region of GABRR2 gene, and both big and small indels can lead to unexpected high probability of exon truncation/skipping. In addition, random deletions in genomic region recruited introns to generate new "exon". It is the first observation of exon recruitment by CRISPR/Cas9-mediated GABRR2 gene editing. The observations may offer a reference for the future gene splicing study. [ABSTRACT FROM AUTHOR]

[Display omitted] • Fe-Kapok has been made by easy hydrothermal method. • Fe-Kapok showed Fe positive impact for As (V) removal. • High and fast As (V) adsorption. • Fe-Kapok could be regenerated and reused several times. Arsenic (As) is one of the most toxic metals for human health. In the present work, kapok fibers were modified using iron Fe(III) to enhance As(V) adsorption and then characterized using Fourier transform infrared spectroscopy (FT-IR) and scanning electron microscopy (SEM) combined with energy dispersive X-ray spectroscopy (EDS). The impacts of pH, dose of adsorbent, contact time, initial concentration, temperature, and coexisting ions on the adsorption of As(V) onto Fe-modified kapok fiber (Fe-kapok) surfaces were investigated. The results showed that the adsorption process fit more to the Langmuir isotherm equation and pseudo second kinetic equation, thus demonstrating that the adsorption was an endothermic, spontaneous, monolayer chemisorption process with some disorder at the solid/solution interface. The adsorption process was controlled by the film diffusion step, and the maximum adsorption capacity of Fe-kapok was calculated to be approximately 367 µg/g. Further study showed that after 8 adsorption/desorption cycles with 1 regeneration cycle, Fe-kapok was still able to remove 45% of the arsenic (100 µg/L) present in the water. A notable negative impact on the removal of As(V) from water was linked to the concentrations of SO 4 2- and PO 4 3- in the water. [ABSTRACT FROM AUTHOR]

Abstract: Recent discovery of biological function of endothelial cell-specific chemotaxic regulator (ECSCR), previously known as endothelial cell-specific molecule 2 (ECSM2), in modulating endothelial cell migration, apoptosis, and angiogenesis, has made it an attractive molecule in vascular research. Thus, identification of splice variants of ECSCR could provide new strategies for better understanding its roles in health and disease. In this study, we performed a series of blast searches on the human EST database with known ECSCR cDNA sequence (Variant 1), and identified additional three splice variants (Variants 2–4). When examining the ECSCR gene in the human genome assemblies, we found a large unknown region between Exons 9 and 11. By PCR amplification and sequencing, we partially mapped Exon 10 within this previously unknown region of the ECSCR gene. Taken together, in addition to previously reported human ECSCR, we identified three novel full-length splice variants potentially encoding different protein isoforms. We further defined a total of twelve exons and nearly all exon–intron boundaries of the gene, of which only eight are annotated in current public databases. Our work provides new information on gene structure and alternative splicing of the human ECSCR, which may imply its functional complexity. This undoubtedly opens new opportunities for future investigation of the biological and pathological significance of these ECSCR splice variants. [Copyright &y& Elsevier]

Parkinson's disease (PD) is a chronic neurodegenerative disorder characterized by dopaminergic neuron loss, which is related to excessive reactive oxygen species (ROS) accumulation. Endogenous peroxiredoxin-2 (Prdx-2) has potent anti-oxidative and anti-apoptotic effects. Proteomics studies revealed plasma levels of Prdx-2 were significantly lower in PD patients than in healthy individuals. For further study of the activation of Prdx-2 and its role in vitro, SH-SY5Y cells and the neurotoxin 1-methyl-4-phenylpyridinium (MPP+) were used to model PD. ROS content, mitochondrial membrane potential, and cell viability were used to assess the effect of MPP+ in SH-SY5Y cells. JC-1 staining was used to determine mitochondrial membrane potential. ROS content was detected using a DCFH-DA kit. Cell viability was measured using the Cell Counting Kit-8 assay. Western blot detected the protein levels of tyrosine hydroxylase (TH), Prdx-2, silent information regulator of transcription 1 (SIRT1), Bax, and Bcl-2. The results showed that MPP+-induced accumulation of ROS, depolarization of mitochondrial membrane potential, and reduction of cell viability occurred in SH-SY5Y cells. In addition, the levels of TH, Prdx-2, and SIRT1 decreased, while the ratios of Bax and Bcl-2 increased. Then, Prdx-2 overexpression in SH-SY5Y cells showed significant protection against MPP+ -induced neuronal toxicity, as evidenced by the decrease in ROS content, increase in cell viability, increase in the level of TH, and decrease in the ratios of Bax and Bcl-2. Meanwhile, SIRT1 levels increase with the level of Prdx-2. This suggests that the protection of Prdx-2 may be related to SIRT1. In conclusion, this study indicated that overexpression of Prdx-2 reduces MPP+-induced toxicity in SH-SY5Y cells and may be mediated by SIRT1. [ABSTRACT FROM AUTHOR]

In this contribution, we demonstrate the synthesis of hydrazone-linked COF-Paper composites (namely COF-Paper1 and COF-Paper2) constructed from two monomers of benzene-1,3,5-tricarbohydrazide (Bth) and dimethoxyterephthal-aldehyde (Dma) or 2,5-dihydroxyterephthalaldehyde (Dha) via in-situ growth method using filter paper as a matrix. Interestingly, 2D COFs paper composites display excellent luminescence in the solid-state, and the as-synthesized COF-Paper1 and COF-Paper2 can be used as turn-off fluorescence sensors to visually detect Fe3+ ions and UO 2 2+ ions by the naked eye, respectively. Moreover, the visible limit of detection for Fe3+ and UO 2 2+ was 5.6 ng cm−2 and 27 ng cm−2, respectively. XPS and FT-IR results reveal that resulting COF-Papers have significant coordination ability toward target metal ions. This effective coordination induces electron transfer from the electron-rich building module to the electron-deficient metal ion, resulting in the fluorescence quenching process. This work not only provides a methodology to grow and assemble 2D COFs into large size ensembles but also paves a new pathway for visual detection of target metal ions. • Two COF-Paper composites have been constructed via in situ growth method. • The resulting 2D COFs paper composites with excellent luminescence properties. • The synthesized COF-Papers can specifically interact with iron ions or uranyl ions. • The COF-Papers can be used as a sensor for visual detection of target metal ions. [ABSTRACT FROM AUTHOR]

A novel fluorescent covalent organic framework containing trimethyl silicon group (COF-TMS) was designed and prepared via the Povarov reaction to be used to detect fluorine ions, on account of the interaction between the silicon and the fluorine ions. Trimethyl silicon, as an electron-donating functional group, was replaced with hydrogen atom and the electronic state of the whole material changed. In this process, the change of COF-TMS structure resulted in the fluorescence color changing from red to blue obviously. Furthermore, the structure and optical property of COF-TMS did not change when there are anions such as H2PO4−, ClO4−, SO42−, CO32−, Cl−, Br−, I−, and NO3− in the solution. Notably, the crystallinity also became particularly high in this process, with perfectly regular hexagonal pore structures being clearly observed in the micron range. This work provides a new idea for the future study of COFs in the field of fluorescence through the combination of COFs structure and fluorescence color transformation. [ABSTRACT FROM AUTHOR]

Food waste and waste paper are the two largest components of municipal solid waste, and many previous studies found that their co-digestion improved methane yield. This study investigated the anaerobic co-digestion of 0–100% food waste with paper food packages on lab-scale with feedstock-to-inoculum (F/I) ratios from 0.5 to 6 (based on volatile solids, VS) and total solids contents from 8% to 28%. In addition to methane yield, volumetric productivity and process economics were also used to evaluate the operating parameters. Results showed that the highest cumulative methane yield of around 530 L/kg-VS feed was obtained from 100% food waste at F/I ratios of 0.5, 1, and 2; the highest volumetric methane productivity of 44.2 L/L work was achieved with 25% food waste at an F/I ratio of 6; and the highest annual net profit was obtained from 75% food waste at an F/I ratio of 2, which also achieved the highest internal rate of return (20.7%) and the shortest payback period (5 years). Addition of waste paper packages to food waste could enhance the digester stability, and it is beneficial to allow certain amount of paper packages and tissues to be collected together with food waste during the initial source separation. [Display omitted] [ABSTRACT FROM AUTHOR]

Store operated calcium entry(SOCE) is known to play a pivotal role in DCs functions including migration, maturation and antigen-presenting ability. Orai1, the major component of SOCE which mainly pairs with Stim1, is surely involved in the regulation of DCs functions. Leptin is recently found to mature DCs, we aim to evaluate the role of Orai1 in leptin-induced dendritic cells(DCs) maturation process and elucidate the mechanism. To this end, Flow cytometry and ELISA were utilized to detect the costimulatory molecule CD86 expression and IL-12 secretion, respectively. Transwell assay was used to examine DCs migration capacity. To evaluate the activity of SOCE, calcium(Ca 2+ ) imaging was performed. Firstly, we confirmed the positive effects of leptin upon SOCE and Orai1 expression in DCs isolated from mouse bone marrow. Secondly, we showed that the effects of leptin on DCs migration and maturation are Orai1 dependent. Moreover, Janus kinase 2(Jak2) silencing inhibited leptin-induced Orai1 expression and influenced DCs functions including migration and maturation as well as IL-12 secretion. In conclusion, our results imply that leptin regulates Orai1 by activating Jak2 signaling pathway, hence facilitating DCs migration and maturation. [ABSTRACT FROM AUTHOR]

Mood disorders are a group of psychiatric conditions that represent leading global disease burdens. Increasing evidence from clinical and preclinical studies supports that innate immune system dysfunction plays an important part in the pathophysiology of mood disorders. P2X7 receptor, belonging to the ligand-gated ion channel P2X subfamily of purinergic P2 receptors for extracellular ATP, is highly expressed in immune cells including microglia in the central nervous system (CNS) and has a vital role in mediating innate immune response. The P2X7 receptor is also important in neuron-glia signalling in the CNS. The gene encoding human P2X7 receptor is located in a locus of susceptibility to mood disorders. In this review, we will discuss the recent progress in understanding the role of the P2X7 receptor in the pathogenesis and development of mood disorders and in discovering CNS-penetrable P2X7 antagonists for potential uses in in vivo imaging to monitor brain inflammation and antidepressant therapeutics. [ABSTRACT FROM AUTHOR]

Although many prokaryotes have glycolysis alternatives, it's considered as the only energy-generating glucose catabolic pathway in eukaryotes. Here, we managed to create a hybrid-glycolysis yeast. Subsequently, we identified an inositol pyrophosphatase encoded by OCA5 that could regulate glycolysis and respiration by adjusting 5-diphosphoinositol 1,2,3,4,6-pentakisphosphate (5-InsP 7) levels. 5-InsP 7 levels could regulate the expression of genes involved in glycolysis and respiration, representing a global mechanism that could sense ATP levels and regulate central carbon metabolism. The hybrid-glycolysis yeast did not produce ethanol during growth under excess glucose and could produce 2.68 g/L free fatty acids, which is the highest reported production in shake flask of Saccharomyces cerevisiae. This study demonstrated the significance of hybrid-glycolysis yeast and determined Oca5 as an inositol pyrophosphatase controlling the balance between glycolysis and respiration, which may shed light on the role of inositol pyrophosphates in regulating eukaryotic metabolism. [Display omitted] • Hybrid of glycolysis with the phosphoketolase pathway made an efficient cell factory • Oca5, identified as an inositol pyrophosphatase, degrading 5-InsP 7 to InsP 6 • InsPs control gene expression involved in glycolysis and respiration to balance fluxes • The mechanism of Oca5 regulating glycolysis and respiration was elaborated Glycolysis is considered as the only energy-generating glucose catabolic pathway in eukaryotes. Qin et al. establish a hybrid-glycolysis yeast that disrupts the Embden-Meyerhof-Parnas glycolysis pathway and introduced components of the phosphoketolase pathway. They further identify Oca5 as an inositol pyrophosphatase controlling the balance between glycolysis and respiration activities. [ABSTRACT FROM AUTHOR]

[Display omitted] • Metal polymeric composites based on pyrimidine-functionalized COFs (covalent organic frameworks) were fabricated. • Fe@COF-Pyr composite possesses good catalytic performance for oxygen evolution reaction. • Au-Pd@COF-Pyr can promote methane oxidation. Functionalized covalent organic frameworks (COFs) materials are used in various fields due to their unique properties. Herein, the reported pyrimidine-functionalized COFs sample named COF-Pyr is selected as the matrix to provide two COFs-based metal composites Fe@COF-Pyr and Au-Pd@COF-Pyr, which are respectively synthesized through metal coordination and in situ reduction reaction. On the one hand, Fe@COF-Pyr used as the electrocatalyst exhibits a good oxygen evolution reaction activity with the overpotential of only 430 mV at a current density of 10 mA cm−2 in the KOH solution (1 M) electrolyte. It is attributed to the large specific surface area (427 m2/g) and abundant catalytic sites of Fe@COF-Pyr, which help to improve the electrochemical performance. On the other hand, Au-Pd@COF-Pyr is an excellent catalyst for methane conversion that the products of the methane oxidation reaction can be obtained under mild reaction conditions. This work not only provides a newfangled notion for the design and preparation of COFs-based metal composites but also broadens the practical application of COFs materials in the domain of catalysis. [ABSTRACT FROM AUTHOR]

Ligand-gated ion channels play a role in mediating fast synaptic transmission for communication between neurons. However, the structural basis for the functional coupling of the binding and pore domains, resulting in channel opening, remains a topic of intense investigation. Here, a series of α7 nicotinic receptor mutants were constructed for expression in cultured mammalian cells, and their single-channel properties were examined using the patch-clamp technique combined with radio ligand binding and the fluorescence staining technique. We demonstrated that the replacement of the four pore-lining residues in the channel domain of the α7 nicotinic receptor with the hydrophilic residue serine prolongs the open-channel lifetime, although the conductance of these mutants decreases. At the coupling interface between the extracellular and transmembrane domains, when the VRW residues in the Cys-loop were substituted with the corresponding residues (i.e., IYN) in the 5-HT 3A receptor, the single-channel activity elicited by acetylcholine is impaired. This effect occurred despite the expression of the mutant receptors on the cell surface and despite the fact that the apparent K d values were much lower than those of the wild-type α7 receptor. When we further lowered the channel-gating barrier of this chimera to enhance the open-channel probability, the loss of function was rescued. Overall, we explored the microscopic mechanisms underlying the interplay between the channel domains and the coupling interface that affect the channel activity, and we generated an allosteric gating model for the α7 receptor. This model shows that the gating machinery and coupling assembly codetermine the single-channel gating kinetics. These results likely apply to all channels in the Cys-loop receptor family. [ABSTRACT FROM AUTHOR]

[Display omitted] • The COF-Tr-BA was prepared for selective adsorption and Li-S battery. • COF-Tr-BA shows excellent selectivity for genistein isomer by chemical interaction. • The introduction of boric acid group helps to improve electrochemical performance. Functional triazine-based covalent organic frameworks (COFs) with boric acid units, COF-Tr-BA, are designed and synthesized via cycloaddition of imine-linked COFs, COF-Tr, with (4-ethynylphenyl) boronic acid. Owing to the chemical interaction between boric acid units and adjacent dihydroxyl groups, COF-Tr-BA is used as the adsorbent for the selective adsorption of genistein isomer 4′6,7-trihydroxyisoflavone, showing an excellent selectivity and great adsorption performance with 5.4 times adsorption capacity (67.5 mg g−1) as high as 4′5,7-trihydroxyisoflavone (12.5 mg g−1). Meanwhile, COF-Tr-BA as the sulfur host for lithium-sulfur (Li-S) battery also exhibits a better electrochemical property, giving an initial discharge capacity up to 1349 mA h g−1 and a substantial capacity of 627 mA h g−1 after 200 cycles at 0.5C. It is attributed to the sulfiphilic interaction between the introduced boric acid groups and polysulfides (PSs), as well as the lithiophilic interaction between N atoms from quinoline and triazine moieties on the backbone of COF-Tr-BA and lithium ions (Li+), thus alleviating the dissolution and reducing the shuttle effect of LiPSs. The boric acid functionalized material COF-Tr-BA with the dual-function for selective adsorption and Li-S battery proposed here lays a significant foundation for multifunctional COFs as a versatile platform in practical application. [ABSTRACT FROM AUTHOR]

[Display omitted] • A new fluorescent covalent organic frameworks was synthesized. • N , O -bidentate ligation was be used to combine difluoroboron. • 50% B-COF exhibited red fluorescence under ultraviolet excitation. The multi-dentate chelate ligands containing N and O atoms have good chelating ability to boron trifluoride and their complexes show good fluorescence properties. In this work, a fluorescent covalent organic framework (COF) with good crystallinity and porosity, 50% B-COF, was successfully synthesized by coordination of starting COFs 50% COF-OH with boron trifluoride diethyl etherate via N, O-bidentate ligation. Imine bonds are highly polarized. The coordination of nitrogen and boron changes the electron cloud distribution of imine bonds, causing the non-fluorescent 50% COF-OH to be transformed into fluorescent 50% B-COF. 50% B-COF exhibited red fluorescence under ultraviolet excitation. This work provides a new strategy for the design of fluorescent COFs. [ABSTRACT FROM AUTHOR]

Abstract: Estrogen has great potential as a therapeutic agent in focal ischemic brain injury. Amino acids as energy resources and neurotransmitters in the central nervous system are crucial for proper neuronal function and excitability. The proton-coupled amino acid transporter PAT1 has clear potential in drug absorption. In this paper, human brain PAT1 was cloned and expressed in Xenopus oocytes. The effects of estradiol on the activity of PAT1 were investigated. Glycine-induced membrane currents mediated by PAT1 were measured using the two-electrode voltage clamp technique. The amplitude of the glycine-elicited current was decreased progressively with increasing concentrations of β-estradiol. A concentration-dependent outwards current of PAT1 was also detected by the presence of β-estradiol. We conclude that estrogen attenuates the activity of PAT1 by directly closing PAT1 channel. Our results may provide an additional mechanism for estrogen on neurotransmission and neuronal metabolism during ischemic injury. [Copyright &y& Elsevier]

Human neuroblastoma SH-SY5Y cells are a widely-used human neuronal cell model in the study of neurodegeneration. A recent study shows that, 1-methyl-4-phenylpyridine ion (MPP), which selectively causes dopaminergic neuronal death leading to Parkinson's disease-like symptoms, can reduce SH-SY5Y cell viability by inducing H2O2 generation and subsequent TRPM2 channel activation. MPP-induced cell death is enhanced by increasing the TRPM2 expression. By contrast, increasing the TRPM2 expression has also been reported to support SH-SY5Y cell survival after exposure to H2O2, leading to the suggestion of a protective role for the TRPM2 channel. To clarify the role of reactive oxygen species (ROS)-induced TRPM2 channel activation in SH-SY5Y cells, we generated a stable SH-SY5Y cell line overexpressing the human TRPM2 channel and examined cell death and cell viability after exposure to H2O2 in the wild-type and TRPM2-overexpressing SH-SY5Y cells. Exposure to H2O2 resulted in concentration-dependent cell death and reduction in cell viability in both cell types. TRPM2 overexpression remarkably augmented H2O2-induced cell death and reduction in cell viability. Furthermore, H2O2-induced cell death in both the wild-type and TRPM2-overexpressing cells was prevented by 2-APB, a TRPM2 inhibitor, and also by PJ34 and DPQ, poly(ADP-ribose) polymerase (PARP) inhibitors. Collectively, our results show that increasing the TRPM2 expression renders SH-SY5Y cells to be more susceptible to ROS-induced cell death and reinforce the notion that the TRPM2 channel plays a critical role in conferring ROS-induced cell death. It is anticipated that SH-SY5Y cells can be useful for better understanding the molecular and signaling mechanisms for ROS-induced TRPM2-mediated neurodegeneration in the pathogenesis of neurodegenerative diseases. [ABSTRACT FROM AUTHOR]