An Activated GOPS‐poly‐L‐Lysine‐ Coated Glass Surface for the Immobilization of 60mer Oligonucleotides

To explore a method for enhancing the immobilization and hybridization efficiency of oligonucleotides on DNA microarrays, conventional protocols of poly‐L‐lysine coating were modified by means of surface chemistry, namely, the slides were prepared by the covalently coupling of poly‐L‐lysine to a glycidoxy‐modified glass surface. The modified slides were then used to print microarrays for the detection of the SARS coronavirus by means of 60mer oligonucleotide probes. The characteristics of the modified slides concerning immobilization efficiency, hybridization dynamics, and probe stripping cycles were determined. The improved surface exhibited high immobilization efficiency, a good quality uniformity, and satisfactory hybridization dynamics. The spotting concentration of 10 μmol/L can meet the requirements of detection; the spots were approximately 170 nm in diameter; the mean fluorescence intensity of the SARS spots were between 3.2 × 10⁴ and 5.0 × 10⁴ after hybridization. Furthermore, the microarrays prepared by this method demonstrated more resistance to consecutive probe stripping cycles. The activated GOPS‐PLL slide could undergo hybridization stripping cycles for at least three cycles, and the highest loss in fluorescence intensity was found to be only 11.9 % after the third hybridization. The modified slides using the above‐mentioned method were superior to those slides treated with conventional approaches, which theoretically agrees with the fact that modification by surface chemistry attaches the DNA covalently firmly to the slides. This protocol may have great promise in the future for application in large‐scale manufacture.     

Direct effects of atmospheric carbon dioxide concentration on whole canopy dark respiration of rice

The purpose of this study was to test for direct inhibition of rice canopy apparent respiration by elevated atmospheric carbon dioxide concentration ([CO₂]) across a range of short‐term air temperature treatments. Rice (cv. IR‐72) was grown in eight naturally sunlit, semiclosed, plant growth chambers at daytime [CO₂] treatments of 350 and 700 μmol mol^?1. Short‐term night‐time air temperature treatments ranged from 21 to 40 °C. Whole canopy respiration, expressed on a ground area basis (R_d), was measured at night by periodically venting the chambers with ambient air. This night‐time chamber venting and resealing procedure produced a range of increasing chamber [CO₂] which we used to test for potential inhibitory effects of rising [CO₂] on R_d. A nitrous oxide leak detection system was used to correct R_d measurements for chamber leakage rate (L) and also to determine if apparent reductions in night‐time R_d with rising [CO₂] could be completely accounted for by L. The L was affected by both CO₂ concentration gradient between the chamber and ambient air and the inherent leakiness of each individual chamber. Nevertheless, after correcting R_d for L, we detected a rapid and reversible, direct inhibition of R_d with rising chamber [CO₂] for air temperatures above 21 °C. This effect was larger for the 350 compared with the 700 μmol mol^?1 daytime [CO₂] treatment and was also increased with increasing short‐term air temperature treatments. However, little difference in R_d was found between the two daytime [CO₂] treatments when night‐time [CO₂] was at the respective daytime [CO₂]. These results suggest that naturally occurring diurnal changes in both ambient [CO₂] and air temperature can affect R_d. Because naturally occurring diurnal changes in both [CO₂] and air temperature can be expected in a future higher CO₂ world, short‐term direct effects of these environmental variables on rice R_d can also be expected.     

Mutagenesis of tyrosine residues within helix VII in subunit I of the cytochrome <Emphasis Type="Italic">cbb</Emphasis><Subscript>3</Subscript> oxidase from <Emphasis Type="Italic">Rhodobacter capsulatus</Emphasis>

The cbb ₃-type oxidases are members of the heme-copper oxidase superfamily, distant by sequence comparisons, but sharing common functional characteristics. The cbb ₃ oxidases are missing an active-site tyrosine residue that is absolutely conserved in all A and B-type heme-copper oxidases. This tyrosine is known to play a critical role in the catalytic mechanisms of A and B-type oxidases. The absence of this tyrosine in the cbb ₃ oxidases raises the possibility that the cbb ₃ oxidases utilize a different catalytic mechanism from that of the other members of the superfamily, or have this conserved residue in different helices. Recently sequence comparisons indicate that, a tyrosine residues that might be analogous to the active-site tyrosine in other oxidases are present in the cbb ₃ oxidases but these tyrosines originates from a different transmembrane helix within the protein. In this research, three conserved tyrosine residues, Y294, Y308 and Y318, in helix VII were substituted for phenylalanine. Y318F mutant in the Rhodobacter capsulatus oxidase resulted in a fully assembled enzyme with nativelike structure and activity, but Y294F mutant is not assembled and have a catalytic activity. On the other hand, Y308F mutant is fully assembled enzyme with nativelike structure, but lacking catalytic activity. This result indicates that Y308 should be crucial in catalytic activity of the cbb ₃ oxidase of R. capsulatus. These findings support the assumption that all of the heme-copper oxidases utilize the same catalytic mechanism and provide a residue originates from different places within the primary sequence for different members of the same superfamily.     

ROLE OF GLUTAMATE DEHYDROGENASE AND GLUTAMINE SYNTHETASE IN CHLORELLA VULGARIS DURING ASSIMILATION OF AMMONIUM WHEN JOINTLY IMMOBILIZED WITH THE MICROALGAE‐GROWTH‐PROMOTING BACTERIUM AZOSPIRILLUM BRASILENSE1

Enzymatic activities of glutamate dehydrogenase (GDH) and glutamine synthetase (GS) participating in the nitrogen metabolism and related ammonium absorption were assayed after the microalga Chlorella vulgaris Beij. was jointly immobilized with the microalgae‐growth‐promoting bacterium Azospirillum brasilense. At initial concentrations of 3, 6, and 10 mg · L^?1 NH₄⁺, joint immobilization enhances growth of C. vulgaris but does not affect ammonium absorption capacity of the microalga. However, at 8 mg · L^?1 NH₄⁺, joint immobilization enhanced ammonium absorption by the microalga without affecting the growth of the microalgal population. Correlations between absorption of ammonium per cell and per culture showed direct (negative and positive) linear correlations between these parameters and microalga populations at 3, 6, and 10 mg · L^?1 NH₄⁺, but not at 8 mg · L^?1 NH₄⁺, where the highest absorption of ammonium occurred. In all cultures, immobilized and jointly immobilized, having the four initial ammonium concentrations, enzymatic activities of Chlorella are affected by A. brasilense. Regardless of the initial concentration of ammonium, GS activity in C. vulgaris was always higher when jointly immobilized and determined on a per‐cell basis. When jointly immobilized, only at an initial concentration of 8 mg · L^?1 NH₄⁺ was GDH activity per cell higher.     

Kinetic Modeling of Amino Acid Oxidation Catalyzed by a New D‐Amino Acid Oxidase from Arthrobacter protophormiae

Amino acid oxidases, which enantiospecifically catalyze the oxidative deamination of either D‐ or L‐amino acids, belong to the class of oxidoreductases functioning with a tightly bound cofactor. This cofactor favors industrial applications of D‐amino acid oxidases (D‐AAO). Hence, the enzyme is very important for the industrial application in the purification and determination of certain amino acids. In developing the enzyme‐catalyzed reaction for large‐scale production, modeling of the reaction kinetics plays an important role. Therefore, the subject of this study was the kinetics of the oxidative deamination, a very complex reaction system, which is catalyzed by D‐AAO from Arthrobacter protophormiae using its natural substrate D‐methionine and the aromatic amino acid 3,4‐dihydroxyphenyl‐D‐alanine (D‐DOPA). The kinetic parameters determined by the measurement of the initial rate and nonlinear regression were verified in batch reactor experiments by comparing calculated and experimental concentration‐time curves. It was found that the enzyme is highly specific towards D‐methionine (K_m = 0.24 mM) and not as specific to D‐DOPA as a substrate (K_m = 9.33 mM). The enzyme activity towards D‐methionine ( = 3.01 U/mL) was approx. seven times higher than towards D‐DOPA ( = 20.01 U/mL). The enzyme exhibited no activity towards L‐methionine and L‐DOPA. Batch and repetitive batch experiments were performed with both substrates in the presence and in the absence of catalase for hydrogen peroxide decomposition. Their comparison made it possible to conclude that hydrogen peroxide has no negative influence on the enzyme activity.     

Purification,Characterization and Antibacterial Activity of l‐amino Acid Oxidase from Cerastes cerastes

Antibiotic resistance presents a real problem in which new antibacterial molecules from natural secretions could be beneficial in the development of new drugs. In this study, Cerastes cerastes venom was investigated for its antibacterial activity against Gram‐positive and Gram‐negative bacteria. The antibacterial activity was evaluated by measuring the halo inhibition and minimum inhibitory concentration (MIC). An l ‐amino acid oxidase (CcLAAO) was purified from this venom using three chromatographic steps; its homogeneity (60 kDa) was confirmed by SDS‐PAGE. LC–MS/MS analysis of CcLAAO showed similarities with other LAAO enzymes from Echis ocellatus and Viridovipera stejnegeri venoms. CcLAAO presents an antibacterial activity against three bacterial strains (Staphylococcus aureus, Methicillin‐resistant S. aureus, and Pseudomonas aeruginosa) with MIC values of 10, 10, and 20 μg/mL, respectively. However, no effect was observed against Escherichia coli and yeast strains. Kinetic parameters of CcLAAO evaluated on l ‐leucine at pH 8.0 and 20°C were K_m = 0.06 mmol and V_max = 164 mmol/min.     

Could rising aquatic carbon dioxide concentrations favour the invasion of elodeids in isoetid‐dominated softwater lakes?

1. During the past century, isoetid vegetation types in softwater lakes have often been invaded by faster‐growing elodeids. In these C‐limited systems, this may be related to rising aquatic CO₂ levels. 2. In a laboratory experiment we tested the growth response of two elodeid species, Myriophyllum alterniflorum and Callitriche hamulata, at four different CO₂ levels, ranging from 20 to 230 μmol L⁻¹. In addition, we tested the effect of the nutrient status of the sediment on the growth of C. hamulata at the different CO₂ levels. 3. Shoot and root growth increased with rising CO₂ availability. Irrespective of sediment type, growth was minimal to negative at the lowest CO₂ treatment level, while becoming positive at CO₂ levels around 40–50 μmol L⁻¹. Substantial growth was only obtained when the macrophytes were growing on mesotrophic sediments. The plants reached close to maximal growth at CO₂ levels of c. 100 μmol L⁻¹. 4. Within this experiment, the growth of C. hamulata at CO₂ levels above 90 μmol L⁻¹ may have been limited by N and P availability in both sediment types. The growth rate of M. alterniflorum did not seem to be limited by N and P availability, most likely due to its much higher relative root production. 5. The experimental results show that neither M. alterniflorum nor C. hamulata is able to invade isoetid‐dominated softwater lakes at very low aquatic CO₂ concentrations. However, if the sediments contain enough nutrients, a rise in aquatic CO₂ could allow the invasion of elodeid species leading to the subsequent disappearance of slow‐growing isoetids.     

The CO2‐concentrating mechanism in the bloom‐forming cyanobacterium Microcystis aeruginosa (Cyanophyceae) and effects of UVB radiation on its operation1

The bloom‐forming cyanobacterium Microcystis aeruginosa (Kütz.) Kütz. 854 was cultured with 1.05 W · m^?2 ultraviolet‐B radiation (UVBR) for 3 h every day, and the CO₂‐concentrating mechanism (CCM) within this species as well as effects of UVBR on its operation were investigated. Microcystis aeruginosa 854 possessed at least three inorganic carbon transport systems and could utilize external HCO₃^? and CO₂ for its photosynthesis. The maximum photosynthetic rate was approximately the same, but the apparent affinity for dissolved inorganic carbon was significantly decreased from 74.7 μmol · L^?1 in the control to 34.7 μmol · L^?1 in UVBR‐treated cells. At 150 μmol · L^?1 KHCO₃ and pH 8.0, Na⁺‐dependent HCO₃^? transport contributed 43.4%–40.2% to the photosynthesis in the control and 34.5%–31.9% in UVBR‐treated cells. However, the contribution of Na⁺‐independent HCO₃^? transport increased from 8.7% in the control to 18.3% in UVBR‐treated cells. The contribution of CO₂‐uptake systems showed little difference: 47.9%–51.0% in the control and 49.8%–47.2% in UVBR‐treated cells. Thus, the rate of total inorganic carbon uptake was only marginally affected, although UVBR had a differential effect on various inorganic carbon transporters. However, the number of carboxysomes in UVBR‐treated cells was significantly decreased compared to that in the control.     

A novel electroconductive interface based on Fe3O4 magnetic nanoparticle and cysteamine functionalized AuNPs: Preparation and application as signal amplification element to minoring of antigen‐antibody immunocomplex and biosensing of prostate cancer

In this study, a novel electroconductive interface was prepared based on Fe₃O₄ magnetic nanoparticle and cysteamine functionalized gold nanoparticle. The engineered interface was used as signal amplification substrate in the electrochemical analysis of antibody‐antigen binding. For this purpose, biotinilated‐anti‐prostate‐specific antigen (PSA) antibody was bioconjugated with iron oxide magnetic nanoparticles (Fe₃O₄) and drop‐casted on the surface of glassy carbon electrode (GCE). Also, secondary antibody (HRP‐Ab2) encapsulated on gold nanoparticles caped by cysteamine was immobilized on the surface of GCE modified electrode. A transmission electron microscopy images shows that a sandwich immunoreaction was done and binding of Ab1 and Ab2 performed successfully. Various parameters of immunoassay, including the loading of magnetic nanoparticles, the amount of gold nanoparticle conjugate, and the immunoreaction time, were optimized. The detection limit of 0.001 μg. L^?1 of PSA was obtained under optimum experimental conditions. It is found that such magneto‐bioassay could be readily used for simultaneous parallel detection of multiple proteins by using multiple inorganic metal nanoparticle tracers and are expected to open new opportunities for early stage diagnosis of cancer in near future.     

NaCS‐PDMDAAC immobilized cultivation of recombinant Dictyostelium discoideum for soluble human Fas ligand production

Dictyostelium discoideum is a promising eukaryotic host for the expression of heterologous proteins requiring post‐translational modifications. However, the dilute nature of D. discoideum cell culture limits applications for high value proteins production. D. discoideum cells, entrapped in sodium cellulose sulfate/poly‐dimethyl‐diallyl‐ammonium chloride (NaCS‐PDMDAAC) capsules were used for biosynthesis of the heterologous protein, soluble human Fas ligand (hFasL). Semi‐continuous cultivations with capsules recycling were carried out in shake flasks. Also, a scaled‐up cultivation of immobilized D. discoideum for hFasL production in a customized vitreous airlift bioreactor was conducted. The results show that NaCS‐PDMDAAC capsules have desirable biophysical properties including biocompatibility with the D. discoideum cells and good mechanical stability throughout the duration of cultivation. A maximum cell density of 2.02 × 10⁷ cells mL^?1 (equivalent to a maximum cell density of 2.22 × 10⁸ cells mL^?1 in capsules) and a hFasL concentration of 130.40 μg L^?1 (equivalent to a hFasL concentration of 1434.40 μg L^?1 in capsules) were obtained in shake flask cultivation with capsules recycling. Also, a maximum cell density of 1.72 × 10⁷cells mL^?1 (equivalent to a maximum cell density of 1.89 × 10⁸ cells mL^?1 in capsules) and a hFasL concentration of 106.10 μg L^?1 (equivalent to a hFasL concentration of 1167.10 μg L^?1 in capsules) were obtained after ～170 h cultivation in the airlift bioreactor (with a working volume of 200 mL in a 315 mL bioreactor). As the article presents a premier work in the application of NaCS‐PDMDAAC immobilized D. discoideum cells for the production of hFasL, more work is required to further optimize the system to generate higher cell densities and hFasL titers for large‐scale applications. © 2014 American Institute of Chemical Engineers Biotechnol. Prog., 31:424–430, 2015     

Steady-State Oxvgen Kinetics of Terminal Oxidases in Trypanosoma mega*

SYNOPSIS. Steady-state oxygen kinetics of Trypanosoma mega reveal the presence of 3 oxidases. These include an oxidase which is sensitive to salicylhydroxamic acid (SHAM) but insensitive to sodium azide. This oxidase could be the L-α glycerophosphate oxidase present in bloodstream trypanosomes. In addition, an oxidase is present which is azide-sensitive but SHAM-insensitive. This oxidase is inhibited by CO and is probably cytochrome aa₃. A 3rd oxidase is insensitive to both azide and SHAM but is inhibited by CO and is possibly cytochrome o. Reciprocal plots of T. mega reveal the presence of 2 oxidases that are inhibited by CO. These results are discussed in the light of previous evidence suggesting the presence of several oxidases and a branched electron transport system in T. mega.     

Grain Boundary Engineering for Achieving High Thermoelectric Performance in n‐Type Skutterudites

Grain or phase boundaries play a critical role in the carrier and phonon transport in bulk thermoelectric materials. Previous investigations about controlling boundaries primarily focused on the reducing grain size or forming nanoinclusions. Herein, liquid phase compaction method is first used to fabricate the Yb‐filled CoSb₃ with excess Sb content, which shows the typical feature of low‐angle grain boundaries with dense dislocation arrays. Seebeck coefficients show a dramatic increase via energy filtering effect through dislocation arrays with little deterioration on the carrier mobility, which significantly enhances the power factor over a broad temperature range with a high room‐temperature value around 47 μW cm^?2 K^?1. Simultaneously, the lattice thermal conductivity could be further suppressed via scattering phonons via dense dislocation scattering. As a result, the highest average figure of merit ZT of ≈1.08 from 300 to 850 K could be realized, comparable to the best reported result of single or triple‐filled Skutterudites. This work clearly points out that low‐angle grain boundaries fabricated by liquid phase compaction method could concurrently optimize the electrical and thermal transport properties leading to an obvious enhancement of both power factor and ZT .     

Legionella pneumophila utilizes a single‐player disulfide‐bond oxidoreductase system to manage disulfide bond formation and isomerization

Legionella pneumophila uses a single homodimeric disulfide bond (DSB) oxidoreductase DsbA2 to catalyze extracytoplasmic protein folding and to correct DSB errors through protein‐disulfide isomerase (PDI) activity. In Escherichia coli, these functions are separated to avoid futile cycling. In L. pneumophila, DsbA2 is maintained as a mixture of disulfides (S‐S) and free thiols (SH), but when expressed in E. coli, only the SH form is observed. We provide evidence to suggest that structural differences in DsbB oxidases (LpDsbB1 and LpDsbB2) and DsbD reductases (LpDsbD1 and LpDsbD2) (compared with E. coli) permit bifunctional activities without creating a futile cycle. LpdsbB1 and LpdsbB2 partially complemented an EcdsbB mutant while neither LpdsbD1 nor LpdsbD2 complemented an EcdsbD mutant unless DsbA2 was also expressed. When the dsb genes of E. coli were replaced with those of L. pneumophila, motility was restored and DsbA2 was present as a mixture of redox forms. A dominant‐negative approach to interfere with DsbA2 function in L. pneumophila determined that DSB oxidase activity was necessary for intracellular multiplication and assembly/function of the Dot/Icm Type IVb secretion system. Our studies show that a single‐player system may escape the futile cycle trap by limiting transfer of reducing equivalents from LpDsbDs to DsbA2.     

Investigating trophic‐level variability in Celtic Sea fish predators

The trophic level (T_L) mean and variance, and the degree of omnivory for five Celtic Sea fish predators were estimated using a database of stomach content records characterized by a high level of taxonomic resolution. The predators occupied a high position in the food web, i.e. 4·75 for Atlantic cod Gadus morhua, 4·44 for haddock Melanogrammus aeglefinus, 4·88 for European hake Merluccius merluccius, 5·00 for megrim Lepidorhombus whiffiagonis and 5·27 for whiting Merlangius merlangus. The level of taxonomic resolution of the prey did not greatly affect mean T_L predator values; an effect on variance was evident, low resolution masking intra‐population variability in T_L. Generalized additive models (GAM) were used to explain the variability of predator T_L caused by environmental variables (International Council for the Exploration of the Sea, ICES, division and season) and predator characteristics (total length, L_T). Significant year, location season and interaction effects were found for some species and with L_T at the scale of ICES subdivision. The species‐specific variability of T_L could be due to spatio‐temporal variations in prey availability and in predator selectivity following ontogenetic changes. Omnivorous fish T_L was less affected by spatio‐temporal variations. In addition, results showed that the omnivory index and T_L variability provide dissimilar information on predator feeding strategy. Combining information on T_L variability and omnivory allowed between within‐individual and between‐individual components contributing to trophic niche width to be separated and the type of generalization of fish predators to be identified.     

High‐level conversion of L‐lysine into 5‐aminovalerate that can be used for nylon 6,5 synthesis

L ‐Lysine is a potential feedstock for the production of bio‐based precursors for engineering plastics. In this study, we developed a microbial process for high‐level conversion of L ‐lysine into 5‐aminovalerate (5AVA) that can be used as a monomer in nylon 6,5 synthesis. Recombinant Escherichia coli WL3110 strain expressing Pseudomonas putida delta‐aminovaleramidase (DavA) and lysine 2‐monooxygenase (DavB) was grown to high density in fed‐batch culture and used as a whole cell catalyst. High‐density E. coli WL3110 expressing DavAB, grown to an optical density at 600 nm (OD₆₀₀) of 30, yielded 36.51 g/L 5AVA from 60 g/L L ‐lysine in 24 h. Doubling the cell density of E. coli WL3110 improved the conversion yield to 47.96 g/L 5AVA from 60 g/L of L ‐lysine in 24 h. 5AVA production was further improved by doubling the L ‐lysine concentration from 60 to 120 g/L. The highest 5AVA titer (90.59 g/L; molar yield 0.942) was obtained from 120 g/L L ‐lysine by E. coli WL3110 cells grown to OD₆₀₀ of 60. Finally, nylon 6,5 was synthesized by bulk polymerization of ?‐caprolactam and δ‐valerolactam prepared from microbially synthesized 5AVA. The hybrid system demonstrated here has promising possibilities for application in the development of industrial bio‐nylon production processes.     

Comparison of Methods for Achieving 24‐Hour Energy Balance in a Whole‐Room Indirect Calorimeter

Objective: To evaluate and compare methods for achieving 24‐hour energy balance in a whole‐room indirect calorimeter. Research Methods and Procedures: Twenty‐four‐hour energy expenditure (EE) for 34 healthy adults (16 women, 18 men) was measured in a calorimeter during a prestudy day and on a subsequent nonconsecutive assessment day (AD). Several methods for estimating EE on the AD using activity factors or regression equations with data available before the AD [anthropometrics, body composition, resting metabolic rate (RMR), sleeping metabolic rate (SMR) on prestudy day, 24‐hour EE on prestudy day] were compared for predictive accuracy. Results: Use of a 24‐hour calorimeter stay gave the smallest mean absolute error (119 ± 16 kcal/d) and smallest single maximum error (361 kcal/d). However, several other methods were only slightly, and not significantly, less accurate (e.g., mean absolute error = 131 ± 17, 140 ± 20, and 141 ± 22 kcal/d and greatest error = 384, 370, and 593 kcal/d for anthropometric, RMR, and SMR regression equations, respectively). Fat‐free mass alone and SMR with a simple activity factor were seen to be less accurate. Discussion: Our results indicate that there may be some improvement in achieving 24‐hour energy balance in a metabolic chamber by using a preceding 24‐hour calorimeter stay; that only slightly less accurate predictions can be obtained using a combination of anthropometric, body composition, and/or RMR measurements; and that there is little or no advantage in using SMR from a previous overnight calorimeter stay.     

Candidacidal mechanism of a Leu/Lys‐rich α‐helical amphipathic model antimicrobial peptide and its diastereomer composed of D,L‐amino acids

We investigated the mechanism of candidacidal action of a Lys/Leu‐rich α‐helical model antimicrobial peptide (K₉L₈W) and its diastereomeric peptide (D₉‐K₉L₈W) composed of D ,L ‐amino acids. K₉L₈W killed completely Candida albicans within 30 min, but D₉‐K₉L₈W killed only 72% of C. albicans even after 100 min. Tryptophan fluorescence spectroscopy indicated that the fungal cell selectivity of D₉‐K₉L₈W is closely correlated with a selective interaction with the negatively charged PC/PE/PI/ergosterol (5:2.5:2.5:1, w/w/w/w) phospholipids, which mimic the outer leaflet of the plasma membrane of C. albicans. K₉L₈W was able to induce almost 100% calcein leakage from PC/PE/PI/ergosterol (5:2.5:2.5:1, w/w/w/w) liposomes at a peptide:lipid molar ratio of 1:16, whereas D₉‐K₉L₈W caused only 25% dye leakage even at a peptide:lipid molar ratio of 1:2. Confocal laser‐scanning microscopy revealed that FITC‐labeled D₉‐K₉L₈W penetrated the cell wall and cell membrane and accumulated inside the cells, whereas FITC‐labeled K₉L₈W did not penetrate but associated with the membranes. Collectively, our results demonstrated that the candidacidal activity of K₉L₈ W and D₉‐K₉L₈W may be due to the transmembrane pore/channel formation or perturbation of the fungal cytoplasmic membranes and the inhibition of intracellular functions, respectively. Finally, D₉‐K₉L₈W with potent anti‐Candida activity but no hemolytic activity may be potentially a useful lead compound for the development of novel antifungal agents. Copyright © 2010 European Peptide Society and John Wiley & Sons, Ltd.     

Production of 1,2‐propanediol in photoautotrophic Synechocystis is linked to glycogen turn‐over

We utilized a photoautotrophic organism to synthesize 1,2‐propanediol from carbon dioxide and water fueled by light. A synthetic pathway comprising mgsA (methylglyoxal synthase), yqhD (aldehyde reductase), and adh (alcohol dehydrogenase) was inserted into Synechocystis sp. PCC6803 to convert dihydroxyacetone phosphate to methylglyoxal, which is subsequently reduced to acetol and then to 1,2‐propanediol. 1,2‐propanediol could be successfully produced by Synechocystis, at an approximate rate of 55 μmol h^?1 g_CDW^?1. Surprisingly, maximal productivity was observed in the stationary phase. The production of 1,2‐propanediol was clearly coupled to the turn‐over of intracellular glycogen. Upon depletion of the glycogen pool, product formation stopped. Reducing the carbon flux to glycogen significantly decreased final product titers. Optimization of cultivation conditions allowed final product titers of almost 1 g L^?1 (12 mM), which belongs to the highest values published so far for photoautotrophic production of this compound.

     

Sub‐6 nm Fully Ordered L10‐Pt–Ni–Co Nanoparticles Enhance Oxygen Reduction via Co Doping Induced Ferromagnetism Enhancement and Optimized Surface Strain

Engineering the crystal structure of Pt–M (M = transition metal) nanoalloys to chemically ordered ones has drawn increasing attention in oxygen reduction reaction (ORR) electrocatalysis due to their high resistance against M etching in acid. Although Pt–Ni alloy nanoparticles (NPs) have demonstrated respectable initial ORR activity in acid, their stability remains a big challenge due to the fast etching of Ni. In this work, sub‐6 nm monodisperse chemically ordered L1₀‐Pt–Ni–Co NPs are synthesized for the first time by employing a bifunctional core/shell Pt/NiCoO_x precursor, which could provide abundant O‐vacancies for facilitated Pt/Ni/Co atom diffusion and prevent NP sintering during thermal annealing. Further, Co doping is found to remarkably enhance the ferromagnetism (room temperature coercivity reaching 2.1 kOe) and the consequent chemical ordering of L1₀‐Pt–Ni NPs. As a result, the best‐performing carbon supported L1₀‐PtNi_0.8Co_0.2 catalyst reveals a half‐wave potential (E_1/2) of 0.951 V versus reversible hydrogen electrode in 0.1 m HClO₄ with 23‐times enhancement in mass activity over the commercial Pt/C catalyst along with much improved stability. Density functional theory (DFT) calculations suggest that the L1₀‐PtNi_0.8Co_0.2 core could tune the surface strain of the Pt shell toward optimized Pt–O binding energy and facilitated reaction rate, thereby improving the ORR electrocatalysis.     

Chemiluminescence of non‐differentiated THP‐1 promonocytes: developing an assay for screening anti‐inflammatory milk proteins and peptides

Human leukemic THP‐1 promonocytes are widely used as a model for peripheral blood monocytes. However, superoxide production during respiratory burst (RB) of non‐differentiated THP‐1 (nd‐THP‐1) cells is very low. Here we present a rapid and low‐cost method for measuring the chemiluminescence (CL) of opsonized zymosan (OZ) induced RB which allows detection of Escherichia coli lipopolysaccharide (LPS) induced priming of nd‐THP‐1 cells on the basis of CL reaction kinetics. Maximum CL intensity obtained was 2.20 ± 0.25 and 1.30 ± 0.11 relative light units, while CL peak time was achieved at 18.1 ± 2.6 and 28.7 ± 1.3 min in primed and non‐primed cells, respectively. The priming of nd‐THP‐1 cells with LPS evoked typical TNF‐α and IL‐6 production. We tested the effects of bovine lactoferrin and protein fractions from Lactobacillus helveticus BGRA43 fermented milk for potential anti‐inflammatory effects on LPS primed nd‐THP‐1 cells. Four fractions were found to inhibit the OZ‐induced CL in a dose‐dependent manner (IC₅₀ 3–30 µg/mL), while lactoferrin inhibited CL to a lesser extent (IC₅₀ 270 µg/mL). These results suggest that measuring CL response of nd‐THP‐1 cells can serve as a method for screening anti‐inflammatory compounds which could be used in reducing the risk of phagocyte‐mediated inflammatory diseases. Copyright © 2010 John Wiley & Sons, Ltd.