Multiple Mono Research Articles

Staphylococcus aureus microbial biofilms degradation using cellobiose dehydrogenase from Thermothelomyces thermophilus M77

This work reports biochemical characterization of Thermothelomyces thermophilus cellobiose dehydrogenase (TthCDHIIa) and its application as an antimicrobial and antibiofilm agent. We demonstrate that TthCDHIIa is thermostable in different ionic solutions and is capable of oxidizing multiple mono and oligosaccharide substrates and to continuously produce H2O2. Kinetics measurements depict the enzyme catalytic characteristics consistent with an Ascomycota class II CDH. Our structural analyses show that TthCDHIIa substrate binding pocket is spacious enough to accommodate larger cello and xylooligosaccharides. We also reveal that TthCDHIIa supplemented with cellobiose reduces the viability of S. aureus ATCC 25923 up to 32 % in a planktonic growth model and also inhibits its biofilm growth on 62.5 %. Furthermore, TthCDHIIa eradicates preformed S. aureus biofilms via H2O2 oxidative degradation of the biofilm matrix, making these bacteria considerably more susceptible to gentamicin and tetracycline.

Multifunctional blazed gratings for multiband spatial filtering, retroreflection, splitting, and demultiplexing based on C2 symmetric photonic crystals

The concept of multifunctional reflection-mode gratings that are based on rod-type photonic crystals (PhCs) with C2 symmetry is introduced. The specific modal properties lead to the vanishing dependence of the first-negative-order maximum on the angle of incidence and the nearly sinusoidal redistribution of the incident-wave energy between zero order (specular reflection) and first negative diffraction order (deflection) at frequency variation. These features are key enablers of diverse functionalities and the merging of different functionalities into one structure. The elementary functionalities, of which multifunctional scenarios can be designed, include but are not restricted to multiband spatial filtering, multiband splitting, retroreflection, and demultiplexing. The proposed structures are capable of multifunctional operation in the case of a single polychromatic incident wave or multiple mono-/polychromatic waves incident at different angles. The generalized demultiplexing is possible in the case of several polychromatic waves. The aforementioned deflection properties yield merging demultiplexing with splitting in one functionality. In turn, it may contribute to more complex multifunctional scenarios. Finally, the proposed PhC gratings are studied in transmissive configuration, in which they show some unusual properties.

Protein dynamics and lipid affinity of monomeric, zeaxanthin-binding LHCII in thylakoid membranes

The xanthophyll cycle in the antenna of photosynthetic organisms under light stress is one of the most well-known processes in photosynthesis, but its role is not well understood. In the xanthophyll cycle, violaxanthin (Vio) is reversibly transformed to zeaxanthin (Zea) that occupies Vio binding sites of light-harvesting antenna proteins. Higher monomer/trimer ratios of the most abundant light-harvesting protein, the light-harvesting complex II (LHCII), usually occur in Zea accumulating membranes and have been observed in plants after prolonged illumination and during high-light acclimation. We present a combined NMR and coarse-grained simulation study on monomeric LHCII from the npq2 mutant that constitutively binds Zea in the Vio binding pocket. LHCII was isolated from 13C-enriched npq2 Chlamydomonas reinhardtii (Cr) cells and reconstituted in thylakoid lipid membranes. NMR results reveal selective changes in the fold and dynamics of npq2 LHCII compared with the trimeric, wild-type and show that npq2 LHCII contains multiple mono- or digalactosyl diacylglycerol lipids (MGDG and DGDG) that are strongly protein bound. Coarse-grained simulations on npq2 LHCII embedded in a thylakoid lipid membrane agree with these observations. The simulations show that LHCII monomers have more extensive lipid contacts than LHCII trimers and that protein-lipid contacts are influenced by Zea. We propose that both monomerization and Zea binding could have a functional role in modulating membrane fluidity and influence the aggregation and conformational dynamics of LHCII with a likely impact on photoprotection ability.

Permeation of Am(III) from Water-Soluble, Organic Polyacids across Hollow-Fiber Renewable Liquid Membranes Facilitated with HDEHP/Dodecane: Contrivance of Chemical Dynamics and Mass Transfer Resistances

An innovatory technique based on the process of selective permeation across the membrane for the treatment of solutions containing polycarboxylic acids strongly bonded with 241Am is formulated. The present article illustrates the parametric results of the process of optimization for the recovery or mitigation of 241Am activity from solutions containing diethylenetriaminepentaacetic acid and lactic acid, applying a methodology of the hollow-fiber renewable liquid membrane (HFRLM). In the liquid-membrane (LM) phase, a cation exchanger-type organic complexant, bis(2-ethylhexyl)hydrophosphoric acid (HDEHP), is incorporated as an ion transporter. The optimized process conditions are as follows: the LM phase containing 0.2 M (HDEHP)2/dodecane, pH of feed solution 1.2 ± 0.05, receiving solution with the LM phase, and 2 M HNO3 (1:4.5) in the form of emulsion. The flow rates of recirculation are maintained as 30 and 20 LPH for solutions of feed and receiving phases, respectively. With an overall mass transfer coefficient, 2.43 × 10–3 cm min–1, 99.9% recovery of Am with its concentration upgradation > 10 times is achieved. The separated product, Am(NO3)3, is amenable for precipitation as Am-oxalate, a precursor to yield AmO2. After HFRLM treatment, the feed solution becomes nonalpha waste for its rightful disposal. For process modeling, the mass transfer coefficient of Am(III) is attributed to the multiple mono- and biphasic chemical equilibria and a series of resistances for ionic diffusion across the HFRLM system. This process has been used successfully to alpha decontaminate, residual solution of the simulated trivalent actinide–lanthanide separation with phosphorus-reagent extraction from aqueous komplex process. In this, the alpha decontamination factor obtained for a single-step operation of 8 h is 836. In the two-step operation of HFRLM, the alpha activity in 5 L residual solution is reduced from 2.34 × 108 to 1.01 × 103 BqL–1.

Biodegradation of PCBs in contaminated water using spent oyster mushroom substrate and a trickle-bed bioreactor

Due to their persistence, polychlorinated biphenyls (PCBs) represent a group of important environmental pollutants, but conventional physicochemical decontamination techniques for their removal are usually expensive. The main aim of this work was to develop a cost-effective method for PCB bioremediation, focusing on contaminated water and utilizing the well-known degradation capability of Pleurotus ostreatus (the oyster mushroom). For this purpose, the conditions of several laboratory-scale reactors (working volume 1 L) were optimized. Spent oyster mushroom substrate obtained from a commercial farm was used as a fungal inoculum and growth substrate. The highest degradation efficiency (87%) was recorded with a continuous low-flow setup, which was subsequently scaled up (working volume 500 L) and used for the treatment of 4000 L of real contaminated groundwater containing 0.1–1 μg/L of PCBs. This trickle-bed pilot-scale bioreactor was able to remove 82, 80, 65, and 30–50% of di-, tri-, tetra- and pentachlorinated PCB congeners, respectively. No degradation was observed for hexa- or heptachlorinated congeners. Multiple mono- and dichlorobenzoic acids (CBAs) were identified as transformation products by mass spectrometry, confirming the role of biodegradation in PCB removal. A Vibrio fischeri bioluminescence inhibition test revealed slight ecotoxicity of the primary reactor effluent (sampling after 24 h), which was quickly suppressed once the effluent passed through the reactor for the second time. Moreover, no other effluent exhibited toxicity for the rest of the experiment (71 days in total). Microbial analyses (phospholipid fatty acid analysis and next-generation sequencing) showed that P. ostreatus was able to degrade PCBs in the presence of an abundance of other fungal species as well as aerobic and anaerobic bacteria. Overall, this study proved the suitability of the use of spent oyster mushroom substrate in a bioremediation practice, even for pollutants as recalcitrant as PCBs.

Divergence in tissue-specific expression patterns of genes associated with the terpeniod biosynthesis in two oregano species Origanum vulgare L., and Origanum majorana

Oregano (Origanum vulgare) and marjoram (O. majorana) are two discrete (in terms of sensory characteristics) species within the genus Origanum (Lamiaceae). Their aroma, flavor, and pharmaceutical value are a result of their essential oils, which comprise mainly of monoterpenes and sesquiterpenes. Marjoram is rich in bicyclic monoterpene cis-sabinene hydrate derived from the biosynthetic “sabinyl” pathway while the phenolic monoterpene carvacrol, arising from the “cymyl” pathway, is the distinctive feature of oregano. To investigate differences between the terpene biosynthetic pathways of both species, we identified key enzymes of terpene biosynthesis from the two species of genus Origanum. The heterologous expression of these enzymes showed that each formed multiple mono- or sesquiterpene products and, in combination, were responsible for the direct production of almost all terpenes found in Origanum essential oils. The correlation between essential oil composition and relative terpene synthase transcript concentrations in O. vulgare and O. majorana demonstrated that monoterpene synthase activity is predominantly regulated at the level of transcription and that the phenolic monoterpene alcohol thymol is derived from α-terpinene, the product of a single monoterpene synthase. The combination of heterologously expressed terpene synthases for in vitro assays resulted in blends of mono- and sesquiterpene products that strongly resembled those found in vivo, indicating that terpene synthase expression levels directly control the composition of the essential oils. These results will facilitate the metabolic engineering, directed breeding of Origanum cultivars with higher quantities of essential oils, and improved oil compositions.

Enhanced reductive dechlorination of trichloroethylene by sulfidated nanoscale zerovalent iron

Direct injection of reactive nanoscale zerovalent iron particles (NZVI) is considered to be a promising approach for remediation of aquifers contaminated by chlorinated organic pollutants. In this study we show that the extent of sulfidation of NZVI enhances the rate of dechlorination of trichloroethylene (TCE) compared to that by unamended NZVI, and the enhancement depends on the Fe/S molar ratio. Experiments where TCE was reacted with NZVI sulfidated to different extents (Fe/S molar ratios 0.62–66) showed that the surface-area normalized first-order TCE degradation rate constant increased up to 40 folds compared to non-sulfidated NZVI. Fe/S ratios in the range of 12–25 provided the highest TCE dechlorination rates, and rates decreased at both higher and lower Fe/S. In contrast, sulfidated NZVI exposed to water in the absence of TCE showed significantly lower hydrogen evolution rate (2.75 μmol L−1 h−1) compared to that by an unamended NZVI (6.92 μmol L−1 h−1), indicating that sulfidation of NZVI suppressed corrosion reactions with water. Sulfide (HS−) ions reacted rapidly with NZVI and X-ray photoelectron spectroscopy analyses showed formation of a surface layer of FeS and FeS2. We propose that more electrons are preferentially conducted from sulfidated NZVI than from unamended NZVI to TCE, likely because of greater binding of TCE on the reactive sites of the iron sulfide outer layer. Resuspending sulfidated NZVI in sulfide-free or sulfide containing solutions altered the TCE degradation rate constants because of changes in the FeS layer thickness. Sulfidated NZVI maintained its high reactivity in the presence of multiple mono and divalent ions and with polyelectrolyte coatings. Thus, sulfide ions in groundwater can significantly alter NZVI reactivity.

Synthesis of macrocyclic organo-peptide hybrids from ribosomal polypeptide precursors via CuAAC-/hydrazide-mediated cyclization.

Macrocyclic peptides have attracted increasing attention as a potential new source of chemical probes and therapeutics. In particular, their conformationally restricted structure combined with a high degree of functional and stereochemical complexity makes them promising scaffolds for targeting biomolecules with high affinity and selectivity. The exploration of this structural class relies on the availability of efficient and versatile methods for the generation of large and diversified libraries of macrocyclic peptide-based molecules. To this end, we have developed a methodology for the synthesis of hybrid organo-peptide macrocycles via the cyclization of ribosomally derived polypeptide sequences with non-peptidic organic linkers. This strategy relies on the chemoselective and bioorthogonal ligation of azide/hydrazide-based "synthetic precursors" with intein-fused polypeptides harboring a side-chain alkyne functionality. This macrocyclization approach was found to proceed with high efficiency across a range of different target peptide sequences spanning 4-12 residues as well as across multiple mono- and diaryl-based synthetic precursors. This versatility combined with the possibility to integrate non-proteinogenic scaffolds into genetically encoded peptide sequences makes this methodology of particularly high value toward the creation and screening of highly diverse libraries of peptide-based macrocycles.

Structural and catalytic differences in the effect of Co and Mo as promoters for Pt-based aqueous phase reforming catalysts

Reaction rates (site time yields, normalized to CO chemisorption sites) and product selectivity were determined for PtCo, PtMo, and Pt supported on multi-walled carbon nanotubes for aqueous phase reforming of glycerol. The bimetallic PtCo and PtMo catalysts had 4.6× and 5.4× higher glycerol consumption rates than Pt, and 3.9× and 0.6× rates in H2 formation compared to Pt. Hydrogen generation selectivity was similar on Pt and PtCo, but PtMo had an increase in selectivity to CO cleavage products, which reduced H2 yield at conversions over 60% (85–90% hydrogen yield for Pt and PtCo, 65% for PtMo). X-ray absorption spectroscopy and scanning transmission electron microscopy results indicate that PtCo adopts multiple mono- and bimetallic structures (Pt shell/Co core, well-mixed alloy, Pt only), which maintain surface Pt sites that are selective to hydrogen generation while adding a promotional metal that increases reaction rates. This is in contrast to a previously characterized PtMo catalyst which introduced surface sites capable of deoxygenation reactions (in the form of PtMo metallic or PtMoO/OH acid pairs) which result in the observed decrease in selectivity.

Linear and branched perfluorooctane sulfonate (PFOS) isomer patterns differ among several tissues and blood of polar bears

Perfluorooctane sulfonate (PFOS) is a globally distributed persistent organic pollutant that has been found to bioaccumulate and biomagnify in aquatic food webs. Although principally in its linear isomeric configuration, 21–35% of the PFOS manufactured via electrochemical fluorination is produced as a branched structural isomer. PFOS isomer patterns were investigated in multiple tissues of polar bears (Ursus maritimus) from East Greenland. The liver (n=9), blood (n=19), brain (n=16), muscle (n=5), and adipose (n=5) were analyzed for linear PFOS (n-PFOS), as well as multiple mono- and di-trifluoromethyl-substituted branched isomers. n-PFOS accounted for 93.0±0.5% of Σ-PFOS isomer concentrations in the liver, whereas the proportion was significantly lower (p<0.05) in the blood (85.4±0.5%). Branched isomers were quantifiable in the liver and blood, but not in the brain, muscle, or adipose. In both the liver and blood, 6-perfluoromethylheptane sulfonate (P6MHpS) was the dominant branched isomer (2.61±0.10%, and 3.26±0.13% of Σ-PFOS concentrations, respectively). No di-trifluoromethyl-substituted isomers were detectable in any of the tissues analyzed. These tissue-specific isomer patterns suggest isomer-specific pharmacokinetics, perhaps due to differences in protein affinities, and thus differences in protein interactions, as well transport, absorption, and/or metabolism in the body.

Influence of Quantum Dot Labels on Single Molecule Movement in the Plasma Membrane

Single particle tracking results are very dependent on the probe that is used. In this study we have investigated the influence that functionalized quantum dots (QDs) have on the recorded movement in single molecule tracking experiments of plasma membrane species in live cells. Potential issues in labeling single molecules with QDs (and other particles e.g. gold particles) are induction of cross-linking of the target molecules, which can cause activation of signaling pathways or reduced mobility, and steric hindrance as a result of the probe size. Cross-linking can be a result of the multivalent functionalization tag (e.g. streptavidin (sAv)) or the presence of multiple mono- or multivalent functionalization tags per QD. In this work, we have compared commercially available sAv-QDs of different sizes with custom prepared Co enzyme A (CoA)-QDs both targeting a GPI-anchored protein modified with either a biotin ligase acceptor peptide (BLAP) or an acyl carrier protein (ACP) tag, respectively. Trajectories of the differently labeled GPI-anchored molecules were recorded simultaneously in dual-color experiments at rates of ∼25 -∼1500 Hz. Knowing the effect of different labels is of utmost importance for simultaneous investigations of different plasma membrane species in order to discriminate the effect of the label from differences in movement of the target molecules.

Terpene synthases of oregano (Origanum vulgare L.) and their roles in the pathway and regulation of terpene biosynthesis

The aroma, flavor and pharmaceutical value of cultivated oregano (Origanum vulgare L.) is a consequence of its essential oil which consists mostly of monoterpenes and sesquiterpenes. To investigate the biosynthetic pathway to oregano terpenes and its regulation, we identified and characterized seven terpene synthases, key enzymes of terpene biosynthesis, from two cultivars of O. vulgare. Heterologous expression of these enzymes showed that each forms multiple mono- or sesquiterpene products and together they are responsible for the direct production of almost all terpenes found in O. vulgare essential oil. The correlation of essential oil composition with relative and absolute terpene synthase transcript concentrations in different lines of O. vulgare demonstrated that monoterpene synthase activity is predominantly regulated on the level of transcription and that the phenolic monoterpene alcohol thymol is derived from gamma-terpinene, a product of a single monoterpene synthase. The combination of heterologously-expressed terpene synthases for in vitro assays resulted in blends of mono- and sesquiterpene products that strongly resemble those found in vivo, indicating that terpene synthase expression levels directly control the composition of the essential oil. These results will facilitate metabolic engineering and directed breeding of O. vulgare cultivars with higher quantity of essential oil and improved oil composition.

Recombinant antibody expression vectors enabling double and triple immunostaining of tissue culture cells using monoclonal antibodies

Next to the already available mouse monoclonal and laboratory animal-derived polyclonal antibodies, recombinant antibodies offer an additional and virtually unlimited arsenal of new immunohistochemical research tools. The major advantages of recombinant antibodies are their rapid and easy generation against virtually any target. The avidity of antibody fragments can be increased by partial dimerisation. This can be achieved by fusion of CL domains derived of different species to recombinant antibody domains. The VL-linker-VH-CL constructs result in significantly lower dimerisation levels compared to the VH-linker-VL-CL antibody constructs. The most efficient dimerisation occurs with the Jun-tagged scFvs. The very large and rapidly expanding collection of recombinant antibodies already available combined with the ease of introducing various tag sequences allows for an almost unrestricted number of easily adjustable research tools. To our best knowledge we report for the first time that using CL domains derived from different species, in combination with readily available commercial secondary antibodies specific for these CL domains, provides an easy method for the application of recombinant monoclonal antibodies of various origins in immunohistochemical analyses eliminating the problem of co-staining with multiple mono- or polyclonal antibodies. Both double and triple labelling experiments can be performed successfully.

Multiple mono- and polymorphic Y-linked copies of the SRY HMG-box in Microtidae

Sex determination in mammals is controlled by SRY (sex-determining region of the Y chromosome), a single-copy gene located on the Y-specific region. Several exceptions to this rule have been described: some rodent species present Y-specific multiple copies (either mono- or polymorphic) of this gene, and two Ellobius species and one Tokudaia species determine sex without a Y chromosome or the SRY gene. Recently, we have described multiple polymorphic copies of the SRY gene in both males and females of the vole species Microtus cabrerae. The female location and the presence of stop codons in some copies from males and females also suggest that they are nonfunctional copies of this gene (pseudogenes). We have investigated the SRY HMG-box in nine species of the family Microtidae; we report here the presence, in eight of these species, of multiple mono- or polymorphic copies of the SRY gene located on the Y chromosome.

Early detection of recurrent mono-metastases of breast cancer to endocrine tissues by following CEA levels

CEA and CA 15-3 are the most commonly used biological markers for detection of recurrences and metastases of breast carcinoma. According to the literature CA 15-3 is the more sensitive of the two markers. Herein, we describe a case of a 50-year old woman with breast cancer, who had multiple mono recurrence of endocrine organ metastases. The CEA levels, rather than CA 15-3, proved to be the more accurate predictor of her disease recurrence. We describe the unique behaviour of her carcinoma, which emphasizes our opinion that the combination of CEA and CA 15-3 could be used to rule out recurrences or metastases of breast cancer.

N-Halogeno compounds Part 19 1: Electrophilic fluorinating agents of the multiple mono- N-fluoro class derived from nitrogen heterocycles

Bis(4-fluoro-1,4-diazoniabicyclo[2.2.2]oct-1-yl)-ethane and -propane tetratriflates ( 4, 5), and 1,1′-difluoro-4,4′-bipyridinium ditriflate ( 6) were prepared by solution-phase direct fluorination of bis(4-aza-1-azoniabicyclo[2.2.2]oct-1-yl)-ethane or -propane ditriflate ( 10,11), and 4,4′-bipyridine respectively. The same technique was employed to convert the complex 4,4′-bipyridine · 2BF 3 to the mono-NF derivative F 3B·NC 5H 4-C 5H 4N +-F BF 4 − ( 7) and its monomethylated analogue (4-NC 5H 4)C 5H 4N +-CH 4 TfO − to F- +NC 5H 4-C 5H 4N +-CH 3 (TfO −) 2 ( 8). The results of site-specific electrophilic fluorination of some model substrates (1-morpholinocyclohexene, phenol, 2-naphthol, anisole, di- and tri-methoxybenzene) with 5,7 and 8 are presented.

Neuropathies périphériques des cryoglobulinémies mixtes essentielles: influence d'une sérologie positive pour le virus de l'hépatite C

Among 15 patients suffering from peripheral neuropathy, 67% had anti-HCV antibodies. The anti-HCV antibodies positive patients had more severe multiple mono- and poly-neuropathies and more frequent inflammatory neuromuscular infiltrates. In some VHC positive cases, the peripheral neuropathy was efficiently treated by interferon alpha.

Levels of Human Chorionic Gonadotropin in Breast Cyst Fluid

The results of dosage of human chorionic gonadotropin (HCG) in 75 breast cyst fluids taken from 61 patients suffering from breast gross cystic disease (BGCD), 13 of which had multiple mono and/or bilateral cysts are discussed. The corresponding sera were also examined. Assays were carried out using the ELISA method. For comparison, HCG levels were also determined in 21 breast cyst fluids using RIA method. In 66.7% of the breast cyst fluids examined the hormone levels were higher than normal serum values. Corresponding sera showed HCG to be within the range of normal levels. In 5 patients the multiple and/or bilateral cysts showed widely differing hormone levels. The importance of these observations is discussed.

Oligoclonal Macroglobulinaemia

Ninety macroglobulinaemia sera were tested for occurrence of multiple mono clonal immunoglobulins. Thirteen (14%) of the sera contained more than one monoclonal immunoglobulin. The homogeneous macroglobulins were isolated from 3 sera; in each case individually specific antigenic determinants common to the two components were demonstrated, in addition to determinants unique for each component. NH2‐terminal amino acid sequence studies in two pa tients showed that the IgM heterogeneity was due to differences in the primary structure of the variable parts of the x (case To) and the μ (case Nae) chains. Macroglobulinaemia appears to be a unique source of homogeneous immuno‐globulins from related clones.