Identification of candidate genes and residues for improving nitrogen use efficiency in the N-sensitive medicinal plant Panax notoginseng.

BACKGROUND: Nitrogen (N) metabolism-related key genes and conserved amino acid sites in key enzymes play a crucial role in improving N use efficiency (NUE) under N stress. However, it is not clearly known about the molecular mechanism of N deficiency-induced improvement of NUE in the N-sensitive rhizomatous medicinal plant Panax notoginseng (Burk.) F. H. Chen. To explore the potential regulatory mechanism, the transcriptome and proteome were analyzed and the three-dimensional (3D) information and molecular docking models of key genes were compared in the roots of P. notoginseng grown under N regimes. RESULTS: Total N uptake and the proportion of N distribution to roots were significantly reduced, but the NUE, N use efficiency in biomass production (NUEb), the recovery of N fertilizer (RNF) and the proportion of N distribution to shoot were increased in the N0-treated (without N addition) plants. The expression of N uptake- and transport-related genes NPF1.2, NRT2.4, NPF8.1, NPF4.6, AVP, proteins AMT and NRT2 were obviously up-regulated in the N0-grown plants. Meanwhile, the expression of CIPK23, PLC2, NLP6, TCP20, and BT1 related to the nitrate signal-sensing and transduction were up-regulated under the N0 condition. Glutamine synthetase (GS) activity was decreased in the N-deficient plants, while the activity of glutamate dehydrogenase (GDH) increased. The expression of genes GS1-1 and GDH1, and proteins GDH1 and GDH2 were up-regulated in the N0-grown plants, there was a significantly positive correlation between the expression of protein GDH1 and of gene GDH1. Glu192, Glu199 and Glu400 in PnGS1 and PnGDH1were the key amino acid residues that affect the NUE and lead to the differences in GDH enzyme activity. The 3D structure, docking model, and residues of Solanum tuberosum and P. notoginseng was similar. CONCLUSIONS: N deficiency might promote the expression of key genes for N uptake (genes NPF8.1, NPF4.6, AMT, AVP and NRT2), transport (NPF1.2 and NRT2.4), assimilation (proteins GS1 and GDH1), signaling and transduction (genes CIPK23, PLC2, NLP6, TCP20, and BT1) to enhance NUE in the rhizomatous species. N deficiency might induce Glu192, Glu199 and Glu400 to improve the biological activity of GS1 and GDH, this has been hypothesized to be the main reason for the enhanced ability of N assimilation in N-deficient rhizomatous species. The key genes and residues involved in improving NUE provide excellent candidates for the breeding of medicinal plants.

The associations of two novel inflammation indexes, systemic immune-inflammation index (SII) and system inflammation response index (SIRI), with periodontitis: evidence from NHANES 2009-2014.

OBJECTIVES: The research's goal is to look for any potential relationships between the systemic immune-inflammation index (SII) and the system inflammation response index (SIRI), along with inflammation indicators and the likelihood of periodontitis. METHODS: Ten thousand two hundred eighty-two individuals in sum were determined to be eligible for this cross-sectional study from the National Health and Nutrition Examination Survey (NHANES) between 2009 and 2014. Multiple logistic regression, generalized additive model, smooth curve fitting, subgroup analysis, and interaction tests were done for analyzing the association between periodontitis and SII, SIRI, and other inflammatory indicators. RESULTS: The analysis, adjusted for population weighting, revealed that individuals with moderate/severe periodontitis had SII levels of 545.46 (95% CI (529.10, 561.82), P = 0.0044) and SIRI levels of 1.33 (95% CI (1.29, 1.37), P < 0.0001). In a fully adjusted multivariate logistic regression model, SII was not sensibly associated with moderate/severe periodontitis among the continuous and quartile Q1-Q4 groups (OR = 0.97, 95% CI (0.91, 1.02)). The continuous variable of SIRI (OR = 1.11, 95% CI (1.06, 1.17)) and the quartile Q4 group (OR = 1.58, 95% CI (1.28, 1.94)) had a deemed significant positive association with moderate to severe periodontitis. In addition, other inflammatory indicators, especially NLR, PPN, PLR, MLR, PC, NC, and MC were observed to be notably involved moderate/severe periodontist in this research. CONCLUSION: We explored the association between periodontitis and two novel comprehensive markers of inflammation (SII and SIRI). CLINICAL RELEVANCE: These inflammatory markers are expected to serve as tools to assist clinicians in diagnosing periodontitis.

Construction of an "ON-OFF" fluoroprobe using ionic liquids-modified orange peel-based carbon quantum dots for selective/sensitive permanganate assay in waters and the underlying quenching mechanisms.

Herein, we fabricated blue-fluorescence carbon quantum dots modified by ionic liquids (ILs-CQDs) with a quantum yield of 18.13% by employing orange peel as a carbon source and [BMIM][H2PO4] as a dopant. The fluorescence intensities (FIs) of ILs-CQDs were significantly quenched upon the addition of MnO4- with excellent selectivity and sensitivity in waters, and this phenomenon provided a feasibility for constructing a sensitive "ON-OFF" fluoroprobe. The prominent overlapping between the maximum excitation/emission of ILs-CQDs and the UV-Vis absorption of MnO4- implied an inner filter effect (IFE). The higher Kq value demonstrated that the fluorescence-quenching phenomenon was a static-quenching process (SQE). Coordination between MnO4- and oxygen/amino-rich groups in ILs-CQDs resulted in the alteration of zeta potential in the fluorescence system. Consequently, the interactions between MnO4- and ILs-CQDs belong to a joint mechanism of IFE and SQE. When plotting the FIs of ILs-CQDs vs. the concentrations of MnO4-, a satisfactorily linear correlation was obtained across the range of 0.3-100 µM with a detectable limit of 0.09 µM. This fluoroprobe was successfully applied to detect MnO4- in environmental waters with satisfactory recoveries of 98.05-103.75% and relative standard deviations (RSDs) of 1.57-2.68%. Also, it gave more excellent performance metrics as compared to the Chinese standard indirect iodometry method and other previous approaches for MnO4- assay. Overall, these findings offer a new avenue to engineer/develop a highly efficient fluoroprobe based on the combination of ILs and biomass-derived CQDs for the rapid/sensitive detection of metal ions in environmental waters.

Glucosylation of Isoeugenol and Monoterpenes in Corynebacterium glutamicum by YdhE from Bacillus lichenformis.

Corynebacterium glutamicum has been regarded as a food-grade microorganism. In recent years, the research to improve the activities of beneficial therapeutics and pharmaceutical substances has resulted in the engineering of the therapeutically favorable cell factory system of C. glutamicum. In this study, we successfully glucosylated isoeugenol and other monoterpene derivatives in C. glutamicum using a promiscuous YdhE, which is a glycosyltransferase from Bacillus lichenformis. For efficient glucosylation, cultivation conditions such as the production time, substrate concentration, carbon source, and culture medium were optimized. Our system successfully converted about 93% of the isoeugenol to glucosylated compounds in the culture. The glucoside compounds were then purified, analyzed, and identified as isoeugenol-1-O-ß-d-glucoside and isoeugenol-1-O-ß-d-(2″-acetyl)-glucoside.

A New Insight into the Capacity Decay Mechanism of Ni-Rich Layered Oxide Cathode for Lithium-Ion Batteries.

Understanding the mapping relationship between electrochemical characteristics and physicochemical properties of layered LiNi0.80 Co0.15 Al0.05 O2 (NCA) cathodes is important to develop high energy density lithium-ion batteries (LIBs). Combining in situ and ex situ characterization, the effect of the H2-H3 phase transition on the capacity decay and aging mechanism of NCA materials are systematically investigated. With the increase of cut-off voltage, the cathode electrolyte interphase (CEI) on the NCA interface shows an evolutionary path of formation-thickening-rupture. This phenomenon is closely related to the H2-H3 phase transition. The volumetric stresses and strains caused by the H2-H3 phase transition accelerate the formation and expansion of secondary particle microcracks in the electrode material, leading to the growth of interfacial CEI variations. The capacity of the electrode material can decrease even if the material does not experience the H2-H3 phase transition due to the persistence of interfacial side reactions with calendar aging from long cycles. This work opens up a valuable perspective for the study of the mapping relationship between phase transition and electrochemical properties in Ni-rich layered oxide cathodes and provides guidance for developing high capacity and long cycle life LIBs.

Frigidibacter oleivorans sp. nov., isolated from a deep well with oil reservoir water.

A Gram-stain-negative, rod-shaped bacterium, designated XJ4T, was isolated from oil-contaminated water, collected from Xinjiang Province, north-west PR China (45° 1' 27″ N, 85° 6' 14″ E). Growth occurred at 20-45 °C (optimum, 30 °C) and pH 6.0-10.0 (optimum, pH 6.0-7.0). Strain XJ4T could tolerate up to 7 % (w/v) NaCl and grow optimally in the absence of NaCl. Phylogenetic analysis based on comparative sequence analysis of 16S rRNA gene sequences indicated that strain XJ4T belonged to the genus Frigidibacter, and that was closely related to Frigidibacter mobilis cai42T (97.2 %), Frigidibacter albus SP32T (97.0 %) and Rhodobacter aestuarii JA296T (97.0 %). The average nucleotide identity values between XJ4T and three type strains were 77.9, 77.6 and 71.9 %, respectively. The DNA G+C content of strain XJ4T was 69.5 mol%. The sole respiratory quinone was Q-10. The major cellular fatty acid was summed feature 8 (C18 : 1 ω7c and/or C18 : 1 ω6c), C18 : 0 and 11-methyl C18 : 1 ω7c. The polar lipids were diphosphatidylglycerol, phosphatidylglycerol, unidentified phospholipids, an unidentified aminolipid and unidentified lipids. On the basis of phenotypic, chemotaxonomic and phylogenetic analyses, strain XJ4T represents a novel species of the genus Frigidibacter, for which the name Frigidibacter oleivorans sp. nov. is proposed. The type strain is XJ4T (=CGMCC 1.13778T=LMG 30952T).

Paracoccus alkanivorans sp. nov., isolated from a deep well with oil reservoir water.

A Gram-stain-negative, non-motile and ovoid bacterial strain, designated 4-2T, was isolated from oil-contaminated water which was collected from Xinjiang Province, north-west PR China. The 16S rRNA gene sequence analysis showed that strain 4-2T belonged to the genus Paracoccus. The species with highest similarity to strain 4-2T was Paracoccus saliphilus YIM 90738T (97.83 %), followed by 'Paracoccus siganidrum' M26 (97.83 %) and Paracoccus endophyticus SYSUP0003T (97.25 %). The average nucleotide identity values between 4-2T and three type strains were 84.69, 77.88 and 74.07 %, respectively. The genomic DNA G+C content of strain 4-2T was 61.4 mol%. Chemotaxonomical characteristic results showed that the respiratory quinone was ubiquinone Q-10 and the major fatty acids were summed feature 8 (C18 : 1 ω7c or C18 : 1 ω6c) and C19 : 0 cyclo ω8c. The polar lipid profile consisted of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylcholine, unidentified phospholipids, an unidentified aminolipid and an unidentified polar lipid. The predominant polyamines were putrescine, cadaverine and spermidine. On the basis of phenotypic, chemotaxonomic and phylogenetic inferences, strain 4-2T represents a novel species of the genus Paracoccus, for which the name Paracoccus alkanivorans sp. nov. is proposed. The type strain is 4-2T (=CGMCC 1.13669T=LMG 30882T).

The coarse-grained models of poly(ethylene oxide) and poly(propylene oxide) homopolymers and poloxamers in big multipole water (BMW) and MARTINI frameworks.

Polyethylene oxide (PEO) and poly(propylene oxide) (PPO), especially their tri-block copolymers PEO-PPO-PEO (poloxamers), have a broad range of applications in biotechnology and medical science. Understanding their specific interactions with biomembranes is the key to unveil the unique features of poloxamers either as membrane-healing or membrane pore-forming agents. Based on the coarse-graining convention of the MARTINI force field and the big multipole water (BMW) model, which has a three charged site topology and can reproduce the correct dipole moment of four-water clusters, we generated coarse-grained (CG) models with analytical and numerical potentials for PEO and PPO homopolymers and poloxamers in dilute solution. The effective bonded interaction potentials between CG beads were determined from the probability distributions of bond lengths, angles and dihedrals that are determined from atomistic simulations. The nonbonded interaction parameters were fine-tuned to reproduce the conformational properties of atomistic PEO and PPO homopolymers and poloxamers via extensive CG simulations of PEO and PPO homopolymers and poloxamers in a BMW water environment. The reported CG models provide a promising framework for a comprehensive understanding of the microstructural, conformational, and dynamic properties of poloxamers and their delicate interactions with other species in an explicit water environment.

Effects of microrna-93 on mouse cardiac microvascular endothelial cells injury and inflammatory response by mediating SPP1 through the NF-ΚB pathway.

BACKGROUND: This study aims to investigate the regulative role of microRNA-93 (miR-93) in mouse cardiac microvascular endothelial cells (CMECs) injury and inflammatory response by negatively targeting SPP1 gene via the NF-κB signaling pathway. METHODS: Healthy Balb/c mice were recruited to establish a mouse model with myocarditis using the CVB3 virus. Mice were grouped into normal, blank, negative control (NC), miR-93 inhibitor, miR-93 mimic, SPP1 short hairpin RNA (shRNA), and miR-93 mimic+SPP1 shRNA groups. Reverse transcription quantitative polymerase chain reaction and Western blot analysis were applied to determine the expressions of miR-93, SPP1, VEGFA, p50, p65, Bax, and Bcl-2. MTT assay was conducted to evaluate cell viability, annexin V-fluorescein isothiocyanate/propidium iodide double staining was conducted to examine cell apoptosis, enzyme-linked immunosorbent assay was conducted to measure secretion of inflammatory factors, and chemical colorimetry was conducted to determine NO secretion. RESULTS: SPP1 was a target gene of miR-93. Compared with the normal group, other six groups showed increased expressions of SPP1, p50, p65, VEGFA, and Bax, as well as cell apoptosis rate and secretion of cell inflammatory factors, and decreased expression of Bcl-2, cell viability, and NO secretion. Compared with the blank group, the miR-93 inhibitor group showed elevated expressions of SPP1, p50, p65, VEGFA, and Bax, as well as cell apoptosis rate and secretion of cell inflammatory factors, and reduced Bcl-2, cell viability, and NO secretion. While the miR-93 mimic and SPP1 shRNA groups displayed opposite results. CONCLUSION: Taking our results together, we conclude that upregulation of miR-93 reduces CMECs injury and inflammatory response by negatively targeting SPP1 via inactivating the NF-κB signaling pathway.

A facile synthesis of perforated reduced graphene oxide for high performance electrochemical sensors.

Highly active perforated reduced graphene oxide (P-rGO) was synthesized by a facile methodology based on co-deposition of graphene oxide with sacrificial Prussian blue. Electrode surface properties were characterized by SEM and EDS. The GC/P-rGO electrode exhibited a larger specific surface area than that of GCE. These findings highlighted that the signal was enhanced for both dopamine detection and selenium detection by using P-rGO as a relevant supporting substrate. The result indicated that the large number of perforated structures formed numerous electrically conductive channels in the structure, improving the electrocatalytic properties.

Evaluation of Mesenteric Lymph Nodes in a Pediatric Population with Mesenteric Lymphadenitis Using Superb Microvascular Imaging.

BACKGROUND This study aimed to evaluate superb microvascular imaging (SMI) as an adjunctive imaging method to evaluate mesenteric lymph nodes in children with mesenteric lymphadenitis compared with healthy children. MATERIAL AND METHODS A retrospective study compared children with mesenteric lymphadenitis (n=27) and healthy children (n=30). Lymph node size was determined using grayscale ultrasonography and parameters of lymph node vascularity were compared using color Doppler flow imaging (CDFI) and SMI. The diagnostic performance of ultrasound (US), US combined with SMI, and US combined with CDFI were compared. RESULTS Lymph nodes from children with mesenteric lymphadenitis (n=77) and normal lymph nodes (n=84) were evaluated by SMI, which showed that the least diameter of lymph nodes in cases of mesenteric lymphadenitis was 0.58±0.15 mm and of normal mesenteric lymph nodes was 0.47±0.08 mm (p<0.001). SMI identified 92.6% of abnormal mesenteric lymph nodes while CDFI detected 85.2%. US combined with SMI had the highest sensitivity (81.5%), and specificity (78.9%) compared with US alone (sensitivity, 63.0%; specificity, 64.9%), and compared with US combined with CDFI (sensitivity, 74.1%; specificity, 75.4%). US combined with SMI and US combined with CDFI achieved the same specificity (76.7%), which was higher than that of US alone (66.7%). CONCLUSIONS SMI was superior to color Doppler flow imaging in evaluating the microvasculature in lymphadenopathy in mesenteric lymphadenitis. SMI may be used as an adjunct to grayscale ultrasonography to assist in identifying mesenteric lymphadenopathy in pediatric patients.

Elevated serum OX40L is a biomarker for identifying corticosteroid resistance in pediatric asthmatic patients.

BACKGROUND: Corticosteroids are widely used to control asthma symptoms, but steroid resistance (SR) is a common adverse reaction. Therefore, it is important to accurately predict the corticosteroid response of asthmatic patients. This study aims to evaluate the serum OX40 ligand (OX40L) in pediatric asthmatic patients, and to investigated its correlations with clinical characteristics and corticosteroid response. METHODS: A total of 192 pediatric asthmatic patients with inhaled corticosteroid (ICS) therapy and 130 healthy controls were selected. Clinical data were collected, and the serum levels of immunoglobulin (IgE), interleukin-6 (IL-6), thymic stromal lymphopoietin (TSLP), and OX40L were measured by enzyme-linked immunosorbent assay (ELISA). The level of serum OX40L was compared between the steroid-sensitive asthma (SSA) and steroid-resistant asthma (SRA) groups. RESULTS: The serum OX40L level in asthmatic patients (713.5 ± 165.7 pg/mL) was significantly higher than that of the healthy controls (238.6 ± 27.8 pg/mL) (P < 0.001), and significantly higher in SRA group (791.2 ± 167.9 pg/mL) than in SSA group (655.6 ± 138.8 pg/mL) (P < 0.001). The serum OX40L level showed a significant positive correlation with serum IgE, blood percentages of eosinophils and neutrophils, serum IL-6 and TSLP, and showed a negative correlation with asthma control test (ACT) score and forced expiratory volume in first second (FEV1%). Receiver operating characteristics (ROC) curve was performed to obtain a cutoff value of serum OX40L as 780 pg/mL (sensitivity = 58.5%; specificity = 86.4%), which can identify SRA in asthmatic patients. Multivariate logistic regression analysis showed that elevated serum OX40L (≥780 pg/mL), as well as lymphocytes (%), ACT score, serum IL-6 and TSLP, were independent predictors of SRA (OX40L ≥ 780 pg/mL: odds ratio = 4.188; 95% CI = 1.800-9.746; P = 0.001). The serum OX40L level was decreased after ICS treatment in asthmatic patients, and the reduction in serum OX40L was significant higher in SSA group compared with SRA group. CONCLUSION: High serum OX40L can be used as a biomarker to identify asthmatic patients with corticosteroid resistance, and the change in OX40L level also reflects the response to ICS treatment. These results suggest an association of OX40L with the pathophysiology, inflammation, and clinical outcomes of asthma. New agents targeting OX40L can provide more precise and personalized therapy for asthma.

Deficiency of unc-51 like kinase 1 (Ulk1) protects against mice traumatic brain injury (TBI) by suppression of p38 and JNK pathway.

Unc-51 like autophagy activating kinase 1 (Ulk1) is a serine/threonine kinase that plays a key role in regulating autophagy processes. We attempted to investigate the effects of Ulk1 on traumatic brain injury (TBI) progression by using wild type (WT) mice and Ulk1-knockout (KO) mice suffered with or not TBI. The results were verified using LPS-treated primary astrocyte (AST). Here, Ulk1 was over-expressed in hippocampus of WT mice after TBI, as well as in lipopolysaccharide (LPS)-stimulated AST. Ulk1-deletion improved cognitive ability and hippocampus histological changes in TBI mice. Nissl and neuronal nuclei (NeuN) staining indicated that Ulk1-deletion increased the number of surviving neurons in hippocampus of TBI mice. Ulk1-ablation alleviated neuroinflammation, as evidenced by the reduced expression of hippocampus pro-inflammatory cytokines in TBI mice. TBI-induced apoptosis was also ameliorated by Ulk1-ablation, as proved by the reduced number of TUNEL-staining cells, and cleaved Caspase-3 and poly (ADP-ribose) polymerase (PARP) expressions. Moreover, Ulk1-knockout suppressed TBI-stimulated activation of astrocytes and microglia cells. Additionally, hippocampus autophagy induced by TBI was attenuated by Ulk1-knockout. Further, TBI-activated p38/c-Jun N-terminal Kinase (JNK) pathway was repressed by Ulk1-deletion in hippocampus of mice. The findings above were confirmed in LPS-stimulated AST with or without Ulk1 siRNA transfection. Intriguingly, pre-treatment of p38 or JNK activator markedly abolished the anti-inflammation, anti-apoptosis and anti-autophagy effects of Ulk1-knockdown on LPS-incubated AST. In conclusion, our results demonstrated that Ulk1 might be a potential target for developing therapeutic strategy against TBI in future.

Various Sequence Types of Escherichia coli Isolates Coharboring blaNDM-5 and mcr-1 Genes from a Commercial Swine Farm in China.

Sixteen different sequence types (STs) of Escherichia coli isolates from a commercial swine farm in China were confirmed to coharbor the carbapenem resistance gene blaNDM-5 and the colistin resistance gene mcr-1 Whole-genome sequencing revealed that blaNDM-5 and mcr-1 were located on a 46-kb IncX3 plasmid and a 32-kb IncX4 plasmid, respectively. The two plasmids can transfer together with a low fitness cost, which might explain the presence of various STs of E. coli coharboring blaNDM-5 and mcr-1.

Dispersal of single and mixed non-albicans Candida species biofilms by ß-1,3-glucanase in vitro.

ß-1,3-glucan plays a role in non-albicans Candida species biofilm formation and survival of biofilm Candida to stresses. In this study, we evaluated the antibiofilm activity of ß-1,3-glucanase, which can degrade poly-ß(1 â†’ 3)-glucose of non-albicans Candida species biofilms, on single and mixed species biofilm of non-albicans Candida species, including Candida tropicalis, Candida parapsilosis and Candida krusei. Biofilm by all tested species in microplate were dispersed more than 60%. ß-1,3-glucanase also detached mixed species biofilm in microplate and on medical material surface. ß-1,3-glucanase had no effect on Candida planktonic growth as well as adhesion. However, further biofilm formation was inhibited with ß-1,3-glucanase added at 24 h after biofilm initiation. ß-1,3-glucanase markedly enhanced the antifungal susceptibility of amphotericin B. The examination using confocal laser scanning microscopy and scanning electron microscope confirmed the antibiofilm activity of ß-1,3-glucanase. Our findings demonstrate that ß-1,3-glucanase may be useful as an antibiofilm agent.

A novel type 1/2 hybrid IncC plasmid carrying fifteen antimicrobial resistance genes recovered from Proteus mirabilis in China.

IncC plasmids are of great concern as vehicles of broad-spectrum cephalosporins and carbapenems resistance genes blaCMY and blaNDM. The aim of this study was to sequence and characterize a multidrug resistance (MDR) IncC plasmid (pPm14C18) recovered from Proteus mirabilis. pPm14C18 was identified in a CMY-2-producing P. mirabilis isolate from chicken in China in 2014, and could be transferred to Escherichia coli conferring an MDR phenotype. Whole genome sequencing confirmed pPm14C18 was a novel type 1/2 hybrid IncC plasmid 165,992bp in size, containing fifteen antimicrobial resistance genes. It harboured a novel MDR mosaic region comprised of a hybrid Tn21tnp-pDUmer, in which blaCTX-M-65, dfrA32 and ereA were firstly reported in IncC plasmid. Phylogenetic relationship reconstruction based on the nucleotide sequences of the 52 IncC backbones showed all type 1 IncC plasmids were clustered into one clade, and then merged with pPm14C18 and finally with the type 2 IncC plasmids and another type 1/2 hybrid IncC plasmid pYR1. The MDR IncC plasmids in P. mirabilis of animal origin might threaten public health, which should be drawn more attention.

Molecular and catalytic properties of fungal extracellular cellobiose dehydrogenase produced in prokaryotic and eukaryotic expression systems.

BACKGROUND: Cellobiose dehydrogenase (CDH) is an extracellular enzyme produced by lignocellulolytic fungi. cdh gene expression is high in cellulose containing media, but relatively low CDH concentrations are found in the supernatant of fungal cultures due to strong binding to cellulose. Therefore, heterologous expression of CDH in Pichia pastoris was employed in the last 15 years, but the obtained enzymes were over glycosylated and had a reduced specific activity. RESULTS: We compare the well-established CDH expression host P. pastoris with the less frequently used hosts Escherichia coli, Aspergillus niger, and Trichoderma reesei. The study evaluates the produced quantity and protein homogeneity of Corynascus thermophilus CDH in the culture supernatants, the purification, and finally compares the enzymes in regard to cofactor loading, glycosylation, catalytic constants and thermostability. CONCLUSIONS: Whereas E. coli could only express the catalytic dehydrogenase domain of CDH, all eukaryotic hosts could express full length CDH including the cytochrome domain. The CDH produced by T. reesei was most similar to the CDH originally isolated from the fungus C. thermophilus in regard to glycosylation, cofactor loading and catalytic constants. Under the tested experimental conditions the fungal expression hosts produce CDH of superior quality and uniformity compared to P. pastoris.

One-Step Synthesis of Water-Dispersible and Biocompatible Silicon Nanoparticles for Selective Heparin Sensing and Cell Imaging.

A sensitive and selective fluorescence "turn-off" sensor to detect heparin using water-soluble silicon nanoparticles (Si NPs) was developed for the first time. The Si NPs were synthesized by a simple one-step procedure, which did not need high-temperature and complex modification. The as-prepared Si NPs featured strong fluorescence, favorable biocompatibility, and robust photo- and pH stability. Significantly, the Si NPs were induced to assemble or aggregate via hydrogen bonding, which resulted in the fluorescence of Si NPs quenched. Under the optimized conditions, the linear range was obtained from 0.02 to 2.0 µg/mL, with a limit of detection of 18 ng/mL (equal to 0.004 U/mL). It was lower than the proper therapeutic level of heparin during cardiovascular surgery and long-term therapy. This proposed method was relatively free of interference from heparin analogues, which commonly existed in heparin samples and could possibly affect heparin detection. Moreover, it did not need to introduce any control medium. As expected, the method was successfully applied to detect heparin in human serum samples with satisfactory recovery ranging from 98.8 to 102.5%. The Si NPs were superbly suitable for cell imaging owing to the negligible cytotoxicity and excellent biocompatibility.

Design of an Os Complex-Modified Hydrogel with Optimized Redox Potential for Biosensors and Biofuel Cells.

Multistep synthesis and electrochemical characterization of an Os complex-modified redox hydrogel exhibiting a redox potential ≈+30 mV (vs. Ag/AgCl 3 M KCl) is demonstrated. The careful selection of bipyridine-based ligands bearing N,N-dimethylamino moieties and an amino-linker for the covalent attachment to the polymer backbone ensures the formation of a stable redox polymer with an envisaged redox potential close to 0 V. Most importantly, the formation of an octahedral N6-coordination sphere around the Os central atoms provides improved stability concomitantly with the low formal potential, a low reorganization energy during the Os(3+/2+) redox conversion and a negligible impact on oxygen reduction. By wiring a variety of enzymes such as pyrroloquinoline quinone (PQQ)-dependent glucose dehydrogenase, flavin adenine dinucleotide (FAD)-dependent glucose dehydrogenase and the FAD-dependent dehydrogenase domain of cellobiose dehydrogenase, low-potential glucose biosensors could be obtained with negligible co-oxidation of common interfering compounds such as uric acid or ascorbic acid. In combination with a bilirubin oxidase-based biocathode, enzymatic biofuel cells with open-circuit voltages of up to 0.54 V were obtained.

Synthetic p55 tandem DNA vaccine against Pneumocystis carinii in rats.

Pneumocystis spp. are opportunistic fungal pathogens that are closely associated with severe pneumonia and pulmonary complications in patients with impaired immunity. In this study, the antigenic epitopes of the gene encoding the 55 kDa antigen fragment of Pneumocystis (p55), which may play an important role in Pneumocystis pneumonia, were analyzed. A gene containing tandem variants of the p55 antigen was synthesized and named the tandem antigen gene (TAG). TAG's potential as a DNA vaccine was assessed in immunosuppressed rats. Immunization with p55-TAG DNA vaccine significantly reduced both the pathogen burden and lung-weight to body-weight ratios. Additionally, p55-TAG vaccination in immunosuppressed rats elicited both cell-mediated and humoral immunity.