Search

Extracellular matrix (ECM) proteins play a pivotal role in cell growth and differentiation. To characterize aged ECM proteins, we compared the proteomes by shotgun method of young (passage #15) and late senescent (passage #40) human dermal fibroblasts (HDFs) using SDS-PAGE coupled with LC–MS/MS. The relative abundance of identified proteins was determined using mol% of individual proteins as a semi-quantitative index. Fifteen ECM proteins including apolipoprotein B (APOB) and high-temperature requirement factor 1 (HTRA1) were up-regulated, whereas 50 proteins including fibronectin 1 (FN1) and vitronectin (VTN) were down-regulated in late senescent HDFs. The identified ECM proteins combined with plasma membrane were queried to construct the protein–protein interaction network using Ingenuity Pathways Analysis, resulting in a distinct FN1-centered network. Of differentially abundant ECM proteins in shotgun proteomics, the protein levels of FN1, VTN, APOB, and HTRA1 were verified by immunoblot analysis. The results suggest that the aging process in HDFs might be finally involved in the impaired FN1 regulatory ECM network combined with altered interaction of neighboring proteins. Shotgun proteomics of highly aged HDFs provides insight for further studies of late senescence-related alterations in ECM proteins. [ABSTRACT FROM AUTHOR]

An amendment to this paper has been published and can be accessed via a link at the top of the paper. [ABSTRACT FROM AUTHOR]

The spiny mouse, Acomys cahirinus displays a unique wound healing ability with regeneration of all skin components in a scar-free manner. To identify orchestrators of this regenerative response we have performed proteomic analyses of skin from Acomys and Mus musculus before and after wounding. Of the ~2000 proteins identified many are expressed at similar levels in Acomys and Mus, but there are significant differences. Following wounding in Mus the complement and coagulation cascades, PPAR signaling pathway and ECM-receptor interactions predominate. In Acomys, other pathways predominate including the Wnt, MAPK, the ribosome, proteasome, endocytosis and tight junction pathways. Notable among Acomys specific proteins are several ubiquitin-associated enzymes and kinases, whereas in Mus immuno-modulation proteins characteristic of inflammatory response are unique or more prominent. ECM proteins such as collagens are more highly expressed in Mus, but likely more important is the higher expression of matrix remodeling proteases in Acomys. Another distinctive difference between Acomys and Mus lies in the macrophage-produced arginase 1 is found in Mus whereas arginase 2 is found in Acomys. Thus, we have identified several avenues for experimental approaches whose aim is to reduce the fibrotic response that the typical mammal displays in response to wounding. [ABSTRACT FROM AUTHOR]

An inter-laboratory study was performed to evaluate the performance of a method developed for the quantification of enrofloxacin in chicken meat. Liquid-liquid extraction combined with a clean-up procedure based on solid-phase extraction followed by a liquid chromatography-tandem mass spectrometric method was used by three individual laboratories. All the investigated results of calibration curves and limits of quantification were within the acceptable range for regulatory testing of enrofloxacin. The three laboratories received blind a certified reference material to analyze in triplicate and assess using statistical analysis. From the results, no statistical differences were found between the laboratories in the precision of the method. Additionally, all the results of the z-score, which is an indication of fixed interval bias criteria for accuracy from the laboratories, fell within the allowable limits (±2σ). Based on this proficiency testing by inter-laboratory comparisons, the analytical method including the sample preparation step was proven to be applicable. [ABSTRACT FROM AUTHOR]

The objective of this study is to demonstrate characteristics of a y-type high molecular weight glutenin subunit (D1y HMW-GS) at Glu-D1 found in IT212991, a North Korean landrace wheat compared to Dy12 and Dy12.K as a novel HMW-GS in JB20, a Korean wheat line onto molecular analyses as PCR, cloning, DNA sequencing, and RP-HPLC and proteomic analyses as sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), two-dimensional electrophoresis (2-DE), Fourier-transform mass spectrometry (LTQ-FT-MS). The D1y of IT212991 was identified to have faster electrophoretic mobility than that of Dy12 by SDS-PAGE. HMW-GS components of IT212991 were identified to be different from Chinese Spring (CS) and JB20, a Korean wheat line by RP-HPLC. The result of mass spectrometric analysis, the D1y of IT212991 (68510.8 Da) was similar to that of Dy12.K of JB20 (68514.4 Da), and lower than Dy12 of CS (69151.2 Da). The result of LTQ-FT-MS based on 2-DE, the D1y of IT212991 was identified to be similar with Dy12 corresponding to the protein function as ‘Glutenin, high molecular weight subunit 12’. The D1y encoding the D1y of IT212991 was identified to consist of 652 amino acid sequences corresponding to 1962 bp according to DNA sequencing. The gene was identified to have a insertion and deletion (InDel) corresponding to 18 bp sequences ‘AACAGGACAAGGGCAACA’ compared to ordinary Dy12 gene. It was demonstrated that the D1y of IT212991 is the same as Dy12.K. [ABSTRACT FROM AUTHOR]

Hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX), a nitroamine explosive, is commonly detected in groundwater at military testing and training sites. The objective of this study was to characterize the microbial community capable of using nitrogen derived from the RDX or RDX intermediates during in situ bioremediation. Active groundwater microorganisms capable of utilizing nitro-, ring- or fully-labeled 15 N-RDX as a nitrogen source were identified using stable isotope probing (SIP) in groundwater microcosms prepared from two wells in an aquifer previously amended with cheese whey to promote RDX biodegradation. A total of fifteen 16S rRNA gene sequences, clustered in Clostridia , β- Proteobacteria , and Spirochaetes , were derived from the 15 N-labeled DNA fractions, suggesting the presence of metabolically active bacteria capable of using RDX and/or RDX intermediates as a nitrogen source. None of the derived sequences matched RDX-degrading cultures commonly studied in the laboratory, but some of these genera have previously been linked to RDX degradation in site groundwater via 13 C-SIP. When additional cheese whey was added to the groundwater samples, 28 sequences grouped into Bacteroidia , Bacilli , and α-, β-, and γ- Proteobacteria were identified. The data suggest that numerous bacteria are capable of incorporating N from ring- and nitro-groups in RDX during anaerobic bioremediation, and that some genera may be involved in both C and N incorporation from RDX. [ABSTRACT FROM AUTHOR]

This study identified microorganisms capable of using the explosive hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) or its metabolites as carbon and/or nitrogen sources under different electron-accepting conditions using 13 C and 15 N stable isotope probing (SIP). Mesocosms were constructed using groundwater and aquifer solids from an RDX-contaminated aquifer. The mesocosms received succinate as a carbon source and one of four electron acceptors (nitrate, manganese(IV), iron(III), or sulfate) or no additional electron acceptor (to stimulate methanogenesis). When RDX degradation was observed, subsamples from each mesocosm were removed and amended with 13 C 3 - or ring- 15 N 3 -, nitro- 15 N 3 -, or fully-labeled 15 N 6 -RDX, followed by additional incubation and isolation of labeled nucleic acids. A total of fifteen 16S rRNA sequences, clustering in α- and γ-Proteobacteria, Clostridia, and Actinobacteria, were detected in the 13 C-DNA fractions. A total of twenty seven sequences were derived from different 15 N-DNA fractions, with the sequences clustered in α- and γ-Proteobacteria, and Clostridia. Interestingly, sequences identified as Desulfosporosinus sp. (in the Clostridia) were not only observed to incorporate the labeled 13 C or 15 N from labeled RDX, but also were detected under each of the different electron-accepting conditions. The data suggest that 13 C- and 15 N-SIP can be used to characterize microbial communities involved in RDX biodegradation, and that the dominant pathway of RDX biodegradation may differ under different electron-accepting conditions. [ABSTRACT FROM AUTHOR]

This study investigated two possible strategies, increasing ammonia oxidation activity and bioaugmenting with triclosan-degrader Sphingopyxis strain KCY1, to enhance triclosan removal in nitrifying activated sludge (NAS). Triclosan (2 mg L −1 ) was removed within 96-h in NAS bioreactors amended with 5, 25 and 75 mg L −1 of ammonium (NH 4 –N). The fastest triclosan removal was observed in 25 mg NH 4 –N L −1 amended-bioreactors where high ammonia oxidation occurred. Inhibition of ammonia oxidation and slower triclosan removal were observed in 75 mg NH 4 –N L −1 amended-bioreactors. Triclosan removal was correlated to the molar ratio of the amount of nitrate produced to the amount of ammonium removed. Bioaugmentation with strain KCY1 did not enhance triclosan removal in the bioreactors with active ammonia oxidation. Approximately 36–42% and 59% of triclosan added were removed within 24-h by ammonia-oxidizing bacteria and unknown triclosan-degrading heterotrophs, respectively. The results suggested that increasing ammonia oxidation activity can be an effective strategy to enhance triclosan removal in NAS. [ABSTRACT FROM AUTHOR]

Abstract: With the availability of plant genome sequencing, analysis of plant proteins with mass spectrometry has become promising and admired. Determining the proteome of a cell is still a challenging assignment, which is convoluted by proteome dynamics and convolution. Chloroplast is fastidious curiosity for plant biologists due to their intricate biochemical pathways for indispensable metabolite functions. In this review, an overview on proteomic studies conducted in wheat with a special focus on subcellular proteomics of chloroplast, salt and water stress. In recent years, we and other groups have attempted to understand the photosynthesis in wheat and abiotic stress under salt imposed and water deficit during vegetative stage. Those studies provide interesting results leading to better understanding of the photosynthesis and identifying the stress-responsive proteins. Indeed, recent studies aimed at resolving the photosynthesis pathway in wheat. Proteomic analysis combining two complementary approaches such as 2-DE and shotgun methods couple to high through put mass spectrometry (LTQ-FTICR and MALDI-TOF/TOF) in order to better understand the responsible proteins in photosynthesis and abiotic stress (salt and water) in wheat chloroplast will be focused. Biological significance: In this review we discussed the identification of the most abundant protein in wheat chloroplast and stress-responsive under salt and water stress in chloroplast of wheat seedlings, thus providing the proteomic view of the events during the development of this seedling under stress conditions. Chloroplast is fastidious curiosity for plant biologists due to their intricate biochemical pathways for indispensable metabolite functions. An overview on proteomic studies conducted in wheat with a special focus on subcellular proteomics of chloroplast, salt and water stress. We have attempted to understand the photosynthesis in wheat and abiotic stress under salt imposed and water deficit during seedling stage. Those studies provide interesting results leading to a better understanding of the photosynthesis and identifying the stress-responsive proteins. In reality, our studies aspired at resolving the photosynthesis pathway in wheat. Proteomic analysis united two complementary approaches such as Tricine SDS-PAGE and 2-DE methods couple to high through put mass spectrometry (LTQ-FTICR and MALDI-TOF/TOF) in order to better understand the responsible proteins in photosynthesis and abiotic stress (salt and water) in wheat chloroplast will be highlighted. This article is part of a Special Issue entitled: Translational Plant Proteomics. [Copyright &y& Elsevier]

Abstract: It is a generally accepted result that the inclusion of rubber particles causes the concrete to degrade physical properties of the concrete. In this study the crumb rubber was partially oxidized and used as additives of mortars. Dramatically, the compressive strength of the rubberized mortars (with 6wt.%) was greater than that of mortars without the crumb rubber. To understand these phenomena, Fourier Transform Infrared Spectroscopy (FTIR) was used to explore the functional groups on surfaces of the crumb rubber, and Scanning Electron Microscopy (SEM) was used to observe microstructures of mortars. With the partial oxidation treatment, rubber surfaces produced hydrophilic functional groups as indicated by IR spectra and the hydration of the cement near the crumb rubber was enhanced as shown by SEM, leading to stronger mortars. [ABSTRACT FROM AUTHOR]

Abstract: Triphenylamine-based novel diamine monomers with side group, such as 4-(2,2′-bithiophenyl)-4′,4″-diaminotriphenylamine (2TTPA) and 4-(3,5-bis(trifluoromehyl)phenyl)-4′,4″-diaminotriphenylamine (6FTPA) were prepared and used for polyimide synthesis. 4-Bromo-dinitrotriphenylamine (BrTPA), prepared from 4-bromoaniline and 1-fluoro-4-nitrobenzene, was reacted with 2,2′-bithiophenene-5-boronic acid (2TBB) or 3,5-bis(trifluoromethyl)benzeneboronic acid (6FBB), followed by hydrogenation to afford 2TTPA and 6FTPA, respectively. After characterization by FT-IR, 1H-NMR, EA and melting point analyzer, the triphenylamine-based dimaines were utilized to prepare polyimides with 3,4,9,10-perylenetetracarboxylic dianhydride (PTCDA) or 2,2-bis-(3,4-dicarboxyphenyl) hexafluoropropane dianhydride (6FDA). The polymers were characterized by FT-IR and 1H-NMR. Thermal, optical and electrical properties were also evaluated by DSC/TGA, UV–vis/photoluminescence spectrometery and cyclic voltammetry (CV), respectively. The 6FDA–6FTPA polyimide exhibited high glass transition temperatures (291°C), high thermal stability (>488°C) and light blue emission (480nm) with very good solubility even after imidization. [ABSTRACT FROM AUTHOR]

Abstract: The International Commission on Radiological Protection (ICRP) published the new recommendations 2007 on radiological protection. International Atomic Energy Agency (IAEA) is also under process in revising its International Basic Safety (BSS). According to revision of the ICRP recommendations and BSS, the Korean government plans to implement those changes. In the 2007 ICRP recommendations, there are some new concepts, principles and quantities. Based on the study carried out by Korea Institute of Nuclear Safety (KINS) so far, the following points are identified as major areas that need further in-depth review and consideration for the implementation of the ICRP 2007 recommendations into Korean radiation protection laws and regulations; changes in the radiation risk factors, radiation weighting factors and tissue weighting factors, maintenance of the ICRP 60 dose limits, practical application of the dose constraints and determination of the reference levels in many source to individual exposure relationships, change from process-based system to exposure situation-based system, strengthening of the principle of optimization in all exposure situations, system of radiation protection for the environment, practical application of the exclusion and exemption principles, active participation of the stakeholders, changes in glossary etc. The study for the implementation of the ICRP 2007 recommendations into national legislations will be conducted till the end of 2012. In the meantime, draft regulations will be developed and the possible impact on the nuclear industry will also be analyzed and active involvement of the stakeholders including licensees will be encouraged in the entire process. The final draft of the revised laws and regulations will be issued in the early of 2013 and the formal legislation process of this final draft will commence in due course. [Copyright &y& Elsevier]

Abstract: Hypoxia during the expansion of adipocytes is known to contribute both to the secretion of multiple inflammation-related adipokines as well as to obesity. We therefore investigated the nature of protein changes occurring in adipocytes during hypoxia by observation of the intracellular proteins that are expressed in 3T3-L1 adipocytes. Lysates were utilized for quantitative proteome analysis using isobaric tags for relative and absolute quantitation (iTRAQ) combined with peptide separation by multi-dimensional liquid chromatography. Antioxidants and elongation factors, as well as glycolytic enzymes were increased in hypoxic adipocytes. These changes were supported by similar changes suggested by real-time PCR. The proteins showing changes are all potential targets for revering the mechanism behind the phenomenon of induction of obese adipocytes by hypoxia. This study can therefore aid in defining the proteomic changes that occur in adipocytes in response to oxygen stress, and can further characterize adipocyte metabolism and adaptation to low oxygen conditions. [Copyright &y& Elsevier]

Acinetobacter lwoffii K24 known as an aniline degrading bacterium has also been found to utilize p-hydroxybenzoate as a sole carbon source. In this study, 2-DE using Q-Sepharose column separation was attempted for fast screening of protocatechuate 3,4-dioxygenase for catabolism of p-hydroxybenzoate in A. lwoffii K24. Two protocatechuate 3,4-dioxygenase subunits, pcaG and pcaH were detected and identified with N-terminal and internal sequencing, suggesting proteomics using a column separation may be helpful for the identification of specific protein spots and maximizing the detectable protein spots on the 2-DE gel. The PCR process using degenerate primers for protocatechuate 3,4-dioxygenase and sequence analyses of the PCR products revealed the existence of pcaH and pcaG in A. lwoffii K24. These two subunits were found to be closely located and share extensive homology with pcaH and pcaG of Pseudomonas marginata or Pseudomonas cepacia, providing the evidence that A. lwoffi K24 has the protocatechuate branches as well as catechol branches of β-ketoadipate pathway. [Copyright &y& Elsevier]

Cav1.2 channel phosphorylation plays an important role in regulating neuronal plasticity by action potential-dependent Ca2+ entry. Most studies of Cav1.2 regulation by phosphorylation have been reported in heart and muscles. Here, we identified phosphorylation sites of neuronal Cav1.2 channel protein purified from rat brain using mass spectrometry. The functional characterization of these phosphorylation sites showed altered voltage-dependent biophysical properties of the channel, without affecting current density. These results show that neuronal Cav1.2 channel is regulated by phosphorylation in a complex mechanism involving multiple phosphorylation sites. [ABSTRACT FROM AUTHOR]

Glechoma hederacea var. longituba (GHL) is one of many herbal plants distributed worldwide and is known to contain various biologically useful antioxidant constituents. GHL has been used in folk remedies for various treatments and as favorable tea beverages. However, research on the precise analysis of ingredients in GHL extracts remains insufficient. In this study, compositional analysis has been conducted on polyphenolic ingredients in GHL hot water extracts. GHL samples collected from growing regions were incubated in hot water at 100 °C for 1 h. The polyphenolic constituents in the hot water extracts were analyzed using high performance liquid chromatography-high resolution mass spectrometry (HPLC-HR MS) and tandem mass spectrometry (HPLC-MS/MS) in negative ion mode. As a result, a total of seven compounds were identified as the major polyphenolic constituents. Interestingly, four constituents out of the identified substances were confirmed to be polyphenol glucuronide conjugates, in which glucuronidation was known to be an important metabolic process in polyphenol aglycone along with methylation and sulphation. This study can be applied for the quality control and standardization of GHL herbal samples and the monitoring of metabolic processes involved in the polyphenolic conjugates. [ABSTRACT FROM AUTHOR]

Rationale: Various medium formulations contain essential fatty acids at concentrations ranging from 10 to 100 mg/L. Accurate and precise lipid measurement in media is crucial for monitoring media quality and conducting studies on lipids in the context of cell culture. This study employed two‐dimensional gas chromatography (GC × GC) analyses to offer enhanced resolution, sensitivity, and separation performance compared to GC. Methods: Quantification of fatty acid methyl esters (FAMEs) in a medium was conducted using GC × GC combined with a high‐resolution mass spectrometer and flame ionization detector, considering potential interference from nonionic surfactant Tween 80, which was precipitated and removed by optimizing the concentration of cobalt thiocyanate (CTA) solution during pretreatment. This advanced analytical approach enabled identification of cis and trans isomers of identical molecular weights and determination of the location and number of double bonds in the same carbon number structure. Results: Our analysis identified 36 FAMEs within the C6–C24 region, and a 5% CTA solution was optimal for efficient removal of Tween 80 during lipid extraction. Additionally, this advanced method minimized FAME contamination and loss during pretreatment, thereby significantly reducing the sample volume required to detect trace levels of FAMEs. This improvement led to a fatty acid recovery rate of 106% while maintaining the average relative standard deviation for the target FAMEs of about 3%. Conclusions: Our research paves the way for future investigation into medium quality control and the role of fatty acids in cell culture. This offers the possibility for economical and effective trace quantification of fatty acids in complex media. [ABSTRACT FROM AUTHOR]

Mass spectrometry (MS)-based intact mass analysis and structural characterization of biotherapeutic proteins such as monoclonal antibodies (mAbs) are a crucial characterization approach from upstream drug development to downstream product analysis. Due to various endogenous modifications leading to the structural heterogeneity and several N-linked glycan species resulting in macro-heterogeneity, it is challenging to characterize the mAbs. Hence, it is essential to understand the micro-heterogeneity of such proteins with high level of complexity which may vary in charge, size, or hydrophobicity. The development of high-throughput native separation techniques hyphenated with MS with high sensitivity and excellent mass accuracy has improved the top/middle down analysis, intact mass detection, subunit analysis, enhanced sequence coverage, and accurate localization of site-specific modifications. In this review, we have focused on the critical inroads taken for the improvement in MS-based techniques to resolve the challenges related to analysis of mAbs. Various MS-based techniques and their role in high-order structural analysis and the progress in software development have been explained, and further, the challenges remaining have been discussed. [ABSTRACT FROM AUTHOR]

Crack formation is a frequent result of residual stress release from colloidal films made by the evaporation of colloidal droplets containing nanoparticles. Crack prevention is a significant task in industrial applications such as painting and inkjet printing with colloidal nanoparticles. Here, we illustrate how colloidal drops evaporate and how crack generation is dependent on the particle size and initial volume fraction, through direct visualization of the individual colloids with confocal laser microscopy. To prevent crack formation, we suggest use of a versatile method to control the colloid-polymer interactions by mixing a nonadsorbing polymer with the colloidal suspension, which is known to drive gelation of the particles with short-range attraction. Gelation-driven crack prevention is a feasible and simple method to obtain crack-free, uniform coatings through drying-mediated assembly of colloidal nanoparticles. [ABSTRACT FROM AUTHOR]

Background: Rice is colonized by plant growth promoting bacteria such as Methylobacterium leading to mutually beneficial plant–microbe interactions. As modulators of the rice developmental process, Methylobacterium influences seed germination, growth, health, and development. However, little is known about the complex molecular responsive mechanisms modulating microbe-driven rice development. The application of proteomics to rice-microbe interactions helps us elucidate dynamic proteomic responses mediating this association. Results: In this study, a total of 3908 proteins were detected across all treatments of which the non-inoculated IR29 and FL478 share up to 88% similar proteins. However, intrinsic differences appear in IR29 and FL478 as evident in the differentially abundant proteins (DAPs) and their associated gene ontology terms (GO). Successful colonization of M. oryzae CBMB20 in rice resulted to dynamic shifts in proteomes of both IR29 and FL478. The GO terms of DAPs for biological process in IR29 shifts in abundance from response to stimulus, cellular amino acid metabolic process, regulation of biological process and translation to cofactor metabolic process (6.31%), translation (5.41%) and photosynthesis (5.41%). FL478 showed a different shift from translation-related to response to stimulus (9%) and organic acid metabolic acid (8%). Both rice genotypes also showed a diversification of GO terms due to the inoculation of M. oryzae CBMB20. Specific proteins such as peptidyl-prolyl cis–trans isomerase (A2WJU9), thiamine thiazole synthase (A2YM28), and alanine—tRNA ligase (B8B4H5) upregulated in IR29 and FL478 indicate key mechanisms of M. oryzae CBMB20 mediated plant growth promotion in rice. Conclusions: Interaction of Methylobacterium oryzae CBMB20 to rice results in a dynamic, similar, and plant genotype-specific proteomic changes supporting associated growth and development. The multifaceted CBMB20 expands the gene ontology terms and increases the abundance of proteins associated with photosynthesis, diverse metabolic processes, protein synthesis and cell differentiation and fate potentially attributed to the growth and development of the host plant. The specific proteins and their functional relevance help us understand how CBMB20 mediate growth and development in their host under normal conditions and potentially link subsequent responses when the host plants are exposed to biotic and abiotic stresses. [ABSTRACT FROM AUTHOR]

Abstract: This study combines Au nanoparticles with TiO2 nanoparticles to form a Schottky barrier, and applies it to the photoelectrode thin film of dye-sensitized solar cells (DSSCs). First, commercial TiO2 powder (Degussa P25) is put in the alkaline solution to prepare for TiO nanotubes (Tnt) by using hydrothermal treatment. Tnt are sintered at 550°C to obtain Tnt-C550 particles. In addition, TiO2 nanoparticles (H180) prepared by subjecting Tnt to two cycles of hydrothermal treatment. H180 served as the first layer, and Tnt-C550 as the second. The third layer adopts salt reduction method to prepare for Au nanoparticles. Experimental results show that the DSSCs prepared with three layers of photoelectrode thin films (H180/Tnt-C550/Au) enhanced the light-to-electricity efficiency of DSSCs by as high as 21% (6.34–7.65%). [Copyright &y& Elsevier]

Sarcopenia, characterized by reduced muscle mass and fiber number leading to muscular atrophy, has been associated with serious socioeconomic challenges among the elderly in developed countries. Therefore, preventing sarcopenia could be a promising strategy for achieving a healthy aging society. Nutmeg (Myristica fragrans) has been used as a spice to increase flavor and prevent putrefaction of food. Nutmeg contains various bioactive components that improve muscle activity. To determine the potential effect of lignan-enriched nutmeg extract (LNX) on sarcopenia, LNX (100 mg/kg body weight)-fed aged mice were subjected to forced exercise. Herein, aged (22-month-old) mice fed LNX for three weeks exhibited a shortened and thickened soleus muscle. The ratio of the soleus muscle mass (%) to body weight was significantly increased in LNX-fed aged mice. The relative increase in muscle mass in LNX-fed aged mice improved exercise activities, including rotarod, swimming, and grip strength test results. Proteome profiles of the soleus muscle of LNX-fed mice were used to analyze protein–protein interaction network. Several myosin heavy chain isoforms were found to interact with actin, ACTA1, which functions as a hub protein. Furthermore, the expression of myogenic proteins, such as MYH1, MYH4, and ACTA1, was dose-dependently increased in vivo. In result, our functional proteomic analysis revealed that feeding LNX restored muscle proteins in aged mice. [ABSTRACT FROM AUTHOR]

This paper describes a simple electrochemical sensing platform based on single-walled carbon nanotube (SWCNT) electrodes for glucose detection. The device fabrication using O2-plasma treatment allows precision and uniformity for the construction of three SWCNT electrodes on the flexible plastic substrate. Glucose assay can be simply accomplished by introducing a glucose sample into the fabricated biosensor. The marked electrocatalytic and biocompatible properties of biosensors based on SWCNT electrodes with the incorporation of ferrocenecarboxylic acid and polyethylene glycol enable effective amperometric measurement of glucose at a low oxidation potential (0.3 V) with low interferences from coexisting species. The device shows efficient electroanalytical performances with high sensitivity (5.5 μA·mM−1·cm−2), good reproducibility (CV less than 3%), and long-term stability (over a month). A linear range of response was found from 0 to 10 mM of glucose with a fast response time of 10 s. This attractive electroanalytical device based on GOx/FCA/PEG/SWCNT electrodes offers a promising system to facilitate a new approach for diverse biosensors and electrochemical devices. [ABSTRACT FROM AUTHOR]

Mutations in PARK7 , the gene encoding the DJ-1 protein, are associated with early onset of Parkinson's disease. The C106 residue of DJ-1 is highly susceptible to oxidation, and its oxidation status is essential for various in vivo neuroprotective roles. Since C106 is readily oxidized to sulfinic acid that is not reduced by dithiothreitol, no method to separate native DJ-1 protein from the oxidized one creates challenges in the in vitro study of the biological relevance of C106-oxidation state. Here, we report an efficient column chromatography method to purify native, C106-sulfinic, and mixed (combination of the priors) forms of DJ-1. This method will be useful for systematic in vitro studies of DJ-1 functions by providing specific native and C106-sulfinic DJ-1 proteins. • DJ-1 is one of the key proteins involved in the early onset of Parkinson's disease. • C106 oxidation (e.g. sulfinic) is crucial for the neuroprotective functions of DJ-1. • Difficulty in the specific management of native and C106-oxidized DJ-1 proteins. • C106 of DJ-1 is highly susceptible to oxidation due to its low pK a value. • Cation-exchange column chromatography separates native and C106-sulfinic forms. [ABSTRACT FROM AUTHOR]

Summary: The omics‐based studies are important for identifying characteristic proteins in plants to elucidate the mechanism of ACC deaminase producing bacteria‐mediated salt tolerance. This study evaluates the changes in the proteome of rice inoculated with ACC deaminase producing bacteria under salt‐stress conditions. Salt stress resulted in a significant decrease in photosynthetic pigments, whereas inoculation of Methylobacterium oryzae CBMB20 had significantly increased pigment contents under normal and salt‐stress conditions. A total of 76, 51 and 33 differentially abundant proteins (DAPs) were identified in non‐inoculated salt‐stressed plants, bacteria‐inoculated plants under normal and salt stress conditions respectively. The abundances of proteins responsible for ethylene emission and programmed cell death were increased, and that of photosynthesis‐related proteins were decreased in non‐inoculated plants under salt stress. However, bacteria‐inoculated plants had shown higher abundance of antioxidant proteins, RuBisCo and ribosomal proteins that are important for enhancing stress tolerance and improving plant physiological traits. Collectively, salt stress might affect plant physiological traits by impairing photosynthetic machinery and accelerating apoptosis leading to a decline in biomass. However, inoculation of plants with bacteria can assist in enhancing photosynthetic activity, antioxidant activities and ethylene regulation related proteins for attenuating salt‐induced apoptosis and sustaining growth and development. [ABSTRACT FROM AUTHOR]

Highlights: [•] Strain PD630 can tolerate low concentration of five model inhibitory compounds. [•] Strain PD630 can use vanillic acid and TPCA for cell growth and TAG accumulation. [•] Vanillin can be used as a carbon source, but TAGs are not accumulated. [•] Strain PD630 can grow on three lignocellulosic hydrolysates and accumulate TAGs. [Copyright &y& Elsevier]

Abstract: Brown adipocytes oxidize fatty acids to produce heat in response to cold or caloric overfeeding. The motivation and function of the development of brown fat may thus counteract obesity, though this remains uncertain. We investigated the brown adipocyte proteome by two-dimensional gel electrophoresis followed by mass spectrometry. Comparative analyses of proteins focused on total protein spots to filter differentially expressed proteins during the differentiation of mouse primary brown preadipocytes. A Western blot analysis was performed to verify the target proteins. The results indicated that 10 protein spots were differentially expressed with significant changes, including the three up-regulated proteins of prohibitin, hypoxanthine–guanine phosphoribosyltransferase, and enoyl-CoA hydratase protein; the 5 down-regulated proteins of triosephosphate isomerase, elongation factor 2, α-tropomyosin slow, endophilin-B1, and cofilin-1 (CFL1); and the two unequivocally expressed proteins of peroxiredoxin-1 and collagen α-1(i) chain precursor. We found that during brown adipogenesis, CFL1 has an inhibitory effect on brown adipocyte differentiation. The overexpression of CFL1 inhibited the brown fat deposition and repressed the brown marker genes UCP1, PRDM16, PGC-1α and PPARγ via actin dynamics and polymerization. These observations may be novel findings that bring new insight into the detailed mechanisms of brown adipogenesis and identify possible therapeutic targets for anti-obesity. Biological significance: We use 2-DE to compare the proteomes of adipocytes during the brown adipogenesis of primary mouse preadipocytes. We identified 10 proteins that are differentially expressed. Among these, we found that the actin binding protein CFL1 inhibits the differentiation of brown preadipocytes. CFL1 overexpressing cells showed lower deposition of brown fat droplets, and the brown marker genes of UCP1, PRDM16, PGC-1α and PPARγ were decreased through actin dynamics and polymerization. [Copyright &y& Elsevier]

Fabrication of a self-packaged partial semicircular nanoporous microchannel is demonstrated using electrospinning, self-limiting ultraviolet lithography, and thermal-reflow packaging processes with SU-8. The fabrication process offers unique advantages of microresolution channel patterning, alignment free seamless integration of nanofibers in a microchannel by self-packaged reflow, and easy formation of 3-D nanoporous channels. A fabricated nanoporous microchannel with a channel length of 7 mm, a channel width of 73.85 $\mu $ m, and a channel height of $34.65~\mu $ m shows slow fluidic flow through the channel, functioning as an effective nanochannel with an equivalent channel height of 800 nm and a radius of 857 nm, and therefore, its reduction ratios of the channel height and radius are $\sim 97.7$ % and 97.8%, respectively. [ABSTRACT FROM AUTHOR]

The fiber of paper mulberry, which is mostly grown in the East Asian region, varies in both length and width—even if from the same species—depending on the country, soil, and climate where it is grown. This accounts for differences in the quality of handmade paper between domestic and imported products. Once handmade paper is manufactured as sheets, its origin cannot be determined from a morphological perspective, regardless of macroscopic and microscopic observations. Therefore, this study attempted to determine the origin of Korean and imported products using mulberry bast fiber and handmade paper from a chemometrics perspective. The inorganic components of wood are absorbed from the soil and exist in the bark of trees; thus, metal and rare-earth elements in the bast fibers and handmade papers were quantitatively analyzed using inductively coupled plasma-atomic emission spectroscopy/mass spectrometry (ICP-AES/MS). Following this, a prediction model of their origin was constructed by applying multivariate statistical analysis, i.e., a partial least squares-discriminant analysis (PLS-DA) loading plot was constructed, which helped in identifying the discrimination factors resulting from their origin. According to the ICP-AES results, Pb was only detected in the Gyeongbuk Mungyeong dak sample of the Korean bast fibers. The Chinese bast fiber sample had high contents of Ca, and B and Zn were detected only in the Chinese bast fiber samples. Bast fiber samples from China, Japan, and Thailand showed relatively high contents of Al, Ca, and Na. Meanwhile, Cu and Zn were detected in the handmade paper manufactured from the imported materials. According to the ICP-MS results, among the Korean handmade papers, the products from Gyeongbuk Mungyeong, Andong, and Chungbuk Goesan exhibited a high U content and that from Gapyeong had high Sc, Rb, Sr, and Ba contents. Thus, the elemental analysis employed in this study effectively identified the origin of the papers. The origin prediction by PLS-DA for the domestic and imported products revealed an accuracy of 86.4% and 72.7% for ICP-AES and ICP-MS methods, respectively, which is deemed acceptable to assess the origin. We believe this method significantly contributes to determining the origin of paper mulberry bast fiber and handmade paper manufactured in East Asia. [ABSTRACT FROM AUTHOR]

Methotrexate (MTX), a folate antagonist, is the anchor drug used to treat several diseases. Therapeutic effects are attributed to intracellular levels of various methotrexate conjugates that are present in the cell as polyglutamates (MTX‐Glu). The present study was conducted to develop a new liquid chromatography‐electrospray ionization‐tandem mass spectrometry (LC‐ESI‐MS/MS)‐based assay to separately quantitate the MTX‐Glu in hair cells, red blood cells, and serum using internal standards. Sample preparation consisted of extraction with an organic solution followed by solid‐phase extraction. The presented methodology was applied for the analysis of methotrexate and its polyglutamates in hair cells, red blood cells, and serum obtained from clinical patients. The developed LC‐ESI‐MS/MS method for the quantitative measurement of MTX‐Glu was both sensitive and precise within the clinically relevant range. This method is possibly be superior with respect to sensitivity, selectivity, and speed than all previously described approaches and can be easily applied in routine clinical tests owing to the combination of a simple pretreatment process with robust LC‐MS/MS. [ABSTRACT FROM AUTHOR]

Trichloroethylene (TCE) is one of the most ubiquitous halogenated organic compounds of concerns of carcinogens in groundwater in Taiwan. Bioremediation has been recognized as a cost-effective approach in reducing TCE concentration. Five pilot-scale wells were constructed to monitor TCE concentrations in contaminated groundwater. With injection of EOS®, TCE was effectively degraded to 42%–93% by the end of 175 days. The biostimulation with EOS® was useful in establishing a micro-site anaerobic but with limited contribution. Dilution of the aquifer movement also caused the TCE reduction among injection and monitoring wells. The degradability was affected by the location and the proximity from the injection well. TCE concentrations found to be negatively correlated with the associated Dehalococcoides spp. and functional genes levels. Dhc concentration of 108 copies L−1 caused the initial 40% of TCE degradation. The well with the optimal degradation owned tceA of 109 cells L−1. T-RFLP results indicate the wells with the superior TCE degradability also performed the highest Shannon index number (means the highest diversity), which occurred on the same day that Dhc levels started to enlarge. Desulfovibrio desulfuricans and Desulfuromonas chloroethenica were predominant species identified in the T-RFLP fingerprint profile. In brief, a variety of different factors including well locations, geochemical indicators, and microbial contribution were useful to explain the site-specific optimal TCE remediation approach. The consistence among TCE degradation, Dhc growing pattern, functional gene levels, and the dynamics of the microbial community structure present the novelty of this study. • TCE degradation achieved approximately 40% when Dehalococcoides sp. levels achieved 108 copies L−1. • T-RFLP results indicate the superior TCE degradability correlated with the highest diversity estimated by H index. • Desulfovibrio desulfuricans and Desulfuromonas chloroethenica were predominant species in the T-RFLP profile. • The TCE degradation of 42%-93% was due to EOS® injection in a micro-site anaerobic condition and dilution of the aquifer. [ABSTRACT FROM AUTHOR]

The Glu- B1al (Bx7 + By8) allele is important for bread-making quality. The allele was found in a Korean wheat landrace using specific DNA markers. Molecular analyses were conducted to identify the overexpressed Bx7 (Bx7) subunit of the allele. The Korean wheat landrace (accession ID: IT166460) showed a similar protein expression level of Bx7 subunit, i.e., overexpression of Bx7 subunit towards cv. Glenlea, Canadian Western Red Spring wheat, which harbors Bx7 subunit of Glu- B1al as detected on SDS-PAGE (sodium dodecyl sulfate poly-acrylamide gel electrophoresis). In addition, 2-DE (two-dimensional electrophoresis) analysis revealed similar protein expression patterns of the Bx7 subunit regions of IT166460 and Glenlea. The proportion of Bx7 to total HMW-GSs (high molecular weight glutenin subunits) in IT166460 (56.17 ± 0.22%) was higher than that of Chinese Spring (34.75 ± 1.03%) and even that of Glenlea (46.25 ± 1.76%) as assessed by RP-HPLC (reverse-phase high-performance liquid chromatography). Overexpression of Bx7 subunit was caused by gene duplication and indels of the promoter region of the Bx7 gene. IT166460 attained the 43 bp indel of the promoter region, as did Glenlea, i.e., the amplicon size of IT166460 was the same as that of Glenlea. In addition, the nucleotides present in the duplicated gene in IT166460 were the same as those in Glenlea. Bx7 subunit is critical for dough strength. However, most wheat accessions harboring the subunit are distributed in America. Furthermore, most Korean wheats have little genetic variation in glutenin composition and are associated with inferior bread quality. Hence, IT166460 could be used to improve bread-making quality in the Korean wheat breeding program. [ABSTRACT FROM AUTHOR]

In this study, we report on the first chemical synthesis of ultra-short pyrazole-arginine based antimicrobial peptidomimetics derived from the newly synthesized N -alkyl/aryl pyrazole amino acids. Through the systematic tuning of hydrophobicity, charge, and peptide length, we identified the shortest peptide Py11 with the most potent antimicrobial activity. Py11 displayed greater antimicrobial activity against antibiotic-resistant bacteria, including MRSA, MDRPA, and VREF, which was approximately 2–4 times higher than that of melittin. Besides its higher selectivity (therapeutic index) toward bacterial cells than LL-37, Py11 showed highly increased proteolytic stability against trypsin digestion and maintained its antimicrobial activity in the presence of physiological salts. Interestingly, Py11 exhibited higher anti-biofilm activity against MDRPA compared to LL-37. The results from fluorescence spectroscopy and transmission electron microscopy (TEM) suggested that Py11 kills bacterial cells possibly by integrity disruption damaging the cell membrane, leading to the cytosol leakage and eventual cell lysis. Furthermore, Py11 displayed significant anti-inflammatory (endotoxin-neutralizing) activity by inhibiting LPS-induced production of nitric oxide (NO) and TNF-α. Collectively, our results suggest that Py11 may serve as a model compound for the design of antimicrobial and antisepsis agents. [ABSTRACT FROM AUTHOR]

Enterococcus faecalis is associated with refractory apical periodontitis and its lipoteichoic acid (Ef.LTA) is considered as a major virulence factor. Although the binding proteins of Ef.LTA may play an important role for mediating infection and immunity in the oral cavity, little is known about Ef.LTA-binding proteins (Ef.LTA-BPs) in saliva. In this study, we identified salivary Ef.LTA-BPs with biotinylated Ef.LTA (Ef.LTA-biotin) through mass spectrometry. The biotinylation of Ef.LTA was confirmed by binding capacity with streptavidin-FITC on CHO/CD14/TLR2 cells. The biological activity of Ef.LTA-biotin was determined based on the induction of nitric oxide and macrophage inflammatory protein-1α in a macrophage cell-line, RAW 264.7. To identify salivary Ef.LTA-BPs, the Ef.LTA-biotin was mixed with a pool of human saliva obtained from nine healthy subjects followed by precipitation with a streptavidin-coated bead. Ef.LTA-BPs were then separated with 12% SDS-PAGE and subjected to the mass spectrometry. Six human salivary Ef.LTA-BPs including short palate lung and nasal epithelium carcinoma-associated protein 2, zymogen granule protein 16 homolog B, hemoglobin subunit α and β, apolipoprotein A-I, and lipocalin-1 were identified with statistical significance ( P < 0.05). Ef.LTA-BPs were validated with lipocalin-1 using pull-down assay. Hemoglobin inhibited the biofilm formation of E. faecalis whereas lipocalin-1 did not show such effect. Collectively, the identified Ef.LTA-BPs could provide clues for our understanding of the pathogenesis of E. faecalis and host immunity in oral cavity. [ABSTRACT FROM AUTHOR]

We have recently shown that a mouse lung infection model resulting in acute pneumonia could be used for evaluating the protective immunity induced by mucosal vaccines against Vibrio cholerae . In order to gain insight and better understanding of the pathogenicity of V. cholerae infection, we identified and compared proteins induced by V. cholerae in nasal washes, bronchoalveolar lavages (BAL), and sera. Intranasal administration of V. cholerae increased the concentration of total proteins in nasal washes and BAL fluids, but not in sera. LTQ-Orbitrap hybrid Fourier transform mass spectrometry showed that cytoskeletal proteins, protease inhibitors and anti-inflammatory mediators were present in nasal washes from uninfected mice. The distinctly expressed proteins in nasal washes in response to V. cholerae mainly consisted of protease inhibitors, anti-inflammatory proteins, and anti-microbial proteins. A number of protease inhibitors and anti-inflammatory proteins were selectively expressed in BAL fluids from V. cholerae -infected mice, while cytoskeletal proteins and heat shock proteins were mainly observed in BAL fluids from uninfected mice. A large number of serum complements, protease inhibitors, and acute phase proteins were expressed in V. cholerae -infected mice. Collectively, these results suggest that intranasal administration of V. cholerae leading to acute pneumonia elicited alterations of protein profiles associated with immune homeostasis and host protection in both the mucosal and systemic compartments. [ABSTRACT FROM AUTHOR]

Peptidoglycan (PGN) is a major cell wall component of Gram-positive bacteria that contributes to the regulation of host immunity in the gastrointestinal tract (GIT). Although Gram-positive bacteria contain structurally distinct PGNs that are considered to differently interact with the GIT, PGN-binding proteins (PGN-BPs) in the GIT have been poorly understood. In the present study, we purified PGNs from Lactobacillus plantarum and Staphylococcus aureus (named as Lp.PGN and Sa.PGN, respectively) and identified Lp.PGN-BPs and Sa.PGN-BPs in the lysate of mouse GIT. Lp.PGN activated nucleotide-binding oligomerization domain (NOD) 1 and NOD2, whereas Sa.PGN activated NOD2, but not NOD1, implying that both PGNs retained the biological activity and were differently recognized by the host cells. PGN-BPs were isolated by precipitation with Lp.PGN or Sa.PGN and identified using LTQ-Orbitrap hybrid Fourier transform mass spectrometry. Three independent experiments demonstrated that 18 Lp.PGN-BPs and 6 Sa.PGN-BPs were reproducibly obtained with statistical significance ( P < 0.05). Both Lp.PGN and Sa.PGN bound to proteins which are related to cytoskeleton, microbial adhesion, and mucosal integrity. Lp.PGN selectively bound to proteins related to gene expression, chaperone, and antimicrobial function. However, Sa.PGN preferentially interacted with proteins involved in adherence and invasion of pathogens. Collectively, these results suggest that bacterial PGNs interact with the proteins regulating mucosal homeostasis and immunity in the gut and PGNs of commensals and pathogens might be also differentially recognized in the GIT. [ABSTRACT FROM AUTHOR]

Helicobacter pylori infection causes a variety of gastrointestinal diseases, including peptic ulcers and gastric cancer. Its colonization of the gastric mucosa of the human stomach is a prerequisite for survival in the stomach. Colonization depends on its motility, which is facilitated by the helical shape of the bacterium. In H. pylori, cross-linking relaxation or trimming of peptidoglycan muropeptides affects the helical cell shape. Csd4 has been identified as one of the cell shape-determining peptidoglycan hydrolases in H. pylori. It is a Zn2+-dependent D,L-carboxypeptidase that cleaves the bond between the γ-D-Glu and the mDAP of the non-cross-linked muramyltripeptide (muramyl-L-Ala-γ-D-Glu- mDAP) of the peptidoglycan to produce the muramyldipeptide (muramyl-L-Ala-γ-D-Glu) and mDAP. Here, the crystal structure of H. pylori Csd4 (HP1075 in strain 26695) is reported in three different states: the ligand-unbound form, the substrate-bound form and the product-bound form. H. pylori Csd4 consists of three domains: an N-terminal D,L-carboxypeptidase domain with a typical carboxypeptidase fold, a central β-barrel domain with a novel fold and a C-terminal immunoglobulin-like domain. The D,L-carboxypeptidase domain recognizes the substrate by interacting primarily with the terminal mDAP moiety of the muramyltripeptide. It undergoes a significant structural change upon binding either mDAP or the mDAP-containing muramyltripeptide. It it also shown that Csd5, another cell-shape determinant in H. pylori, is capable of interacting not only with H. pylori Csd4 but also with the dipeptide product of the reaction catalyzed by Csd4. [ABSTRACT FROM AUTHOR]

Mulitpotent mesenchymal stem cells (MSCs) derived from human bone marrow are promising candidates for the development of cell therapeutic strategies. MSC surface protein profiles provide novel biological knowledge concerning the proliferation and differentiation of these cells, including the potential for identifying therapeutic targets. Basic fibroblast growth factor (bFGF) affects cell surface proteins, which are associated with increased growth rate, differentiation potential, as well as morphological changes of MSCs in vitro. Cell surface proteins were isolated using a biotinylation-mediated method and identified using a combination of one-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis and mass spectrometry. The resulting gel lines were cut into 20 bands and digested with trypsin. Each tryptic fragment was analyzed by liquid chromatography-electrospray ionization tandem mass spectrometry. Proteins were identified using the Mascot search program and the International Protein Index human database. Noble MSC surface proteins ( n = 1,001) were identified from cells cultured either with ( n = 857) or without ( n = 667) bFGF-containing medium in three independent experiments. The proteins were classified using FatiGO to elucidate their function. We also confirmed the proteomics results using Western blotting and immunofluorescence microscopic analysis. The nature of the proteins identified makes it clear that MSCs express a wide variety of signaling molecules, including those related to cell differentiation. Among the latter proteins, four Ras-related Rab proteins, laminin-R, and three 14-3-3 proteins that were fractionated from MSCs cultured on bFGF-containing medium are implicated in bFGF-induced signal transduction of MSCs. Consequently, these finding provide insight into the understanding of the surface proteome of human MSCs. [Figure not available: see fulltext.] [ABSTRACT FROM AUTHOR]

The positioning of the nucleosome by ATP-dependent remodellers provides the fundamental chromatin environment for the regulation of diverse cellular processes acting on the underlying DNA. Recently, genome-wide nucleosome mapping has revealed more detailed information on the chromatin-remodelling factors. Here, we report that the Schizosaccharomyces pombe CHD remodeller, Hrp3, is a global regulator that drives proper nucleosome positioning and nucleosome stability. The loss of Hrp3 resulted in nucleosome perturbation across the chromosome, and the production of antisense transcripts in the hrp3? cells emphasized the importance of nucleosome architecture for proper transcription. Notably, perturbation of the nucleosome in hrp3 deletion mutant was also associated with destabilization of the DNA-histone interaction and cell cycle-dependent alleviation of heterochromatin silencing. Furthermore, the effect of Hrp3 in the pericentric region was found to be accomplished via a physical interaction with Swi6, and appeared to cooperate with other heterochromatin factors for gene silencing. Taken together, our data indicate that a well-positioned nucleosome by Hrp3 is important for the spatial-temporal control of transcription-associated processes. [ABSTRACT FROM AUTHOR]

Abstract: This study investigates a solar-thermoelectric module for power generation from solar energy. The proposed method uses recycled external exhaust heat to generate electric power, further enhancing the thermoelectric conversion efficiency of the thermoelectric generator (TEG). Using electrophoresis deposition, self-prepared CuO nanofluid is deposited onto a Cu plate and then adheres to the surface of a thermoelectric generator (TEG). Experimental results show that the CuO thin film coating on the TEG surface can elevate the temperature by around 2 °C and the voltage by around 14.8%, thus enhancing the thermoelectric conversion efficiency of the thermoelectric generator by 10% and increasing the overall power output by 2.35%. It was found that this solar-thermoelectric module can generate about 4.95 mW/cm2 under solar radiation intensity of about 100 mW/cm2. [Copyright &y& Elsevier]

Abstract: Porphyromonas gingivalis causes periodontal diseases and its lipopolysaccharide (LPS) is considered as a major virulence factor responsible for pathogenesis. Since initial recognition of P. gingivalis LPS (Pg.LPS) in the oral cavity might be crucial for the host response, we identified Pg.LPS-binding proteins (Pg.LPS-BPs) using Pg.LPS-immobilized beads and a high-resolution mass spectrometry. LPS purified from P. gingivalis was conjugated onto N-hydroxysuccinimidyl-Sepharose® 4 Fast Flow beads. Notably, Pg.LPS-conjugated beads could stimulate Toll-like receptor 2 (TLR2) as determined by a TLR2-depdendent reporter expression system using CHO/CD14/TLR2. In addition, the Pg.LPS-conjugated beads induced the production of inflammatory mediators such as nitric oxide and interferon-gamma-inducible protein-10 in the macrophage cell-line, RAW 264.7. These results imply that Pg.LPS retained its immunological properties during the conjugation process. Then, the Pg.LPS-conjugated beads were mixed with a pool of saliva obtained from nine human subjects to capture Pg.LPS-BPs and molecular identities were determined by LTQ-Orbitrap hybrid fourier transform mass spectrometry. Pg.LPS-BPs captured at high frequencies included alpha-amylase, cystatin, prolactin-inducible protein, lysozyme C, immunoglobulin components, serum albumin, lipocalin-1, and submaxillary gland androgen-regulated protein 3B. These proteins are known to be involved in bacterial adhesion and colonization, anti-microbial functions or modulation of immune responses. [Copyright &y& Elsevier]

Abstract: This study attempted to validate a new method for patient-tailored efficiency calibration. Digital calibration with Monte Carlo simulations was used to substitute the lack of precision limitation due to the limited number of experimental phantoms in whole body counting calibration for internal dosimetry. The validity of this approach was examined by comparing the simulation results to the measured values from actual measurements using family BOMAB phantoms. The computational voxel phantoms were constructed by a reconstruction technique using AP and lateral photographic images of the BOMAB phantoms, instead of using the given specifications provided with BOMAB phantoms. Although discrepancies to a certain degree between the computational simulation and measured efficiencies do exist, the results support the new approach of being an alternative to family BOMAB phantoms. [Copyright &y& Elsevier]

Abstract: The identification of membrane proteins is currently under-represented since the trans-membrane domains of membrane proteins have a hydrophobic property. Membrane proteins have mainly been analyzed by cleaving and identifying exposed hydrophilic domains. We developed the membrane proteomics method for targeting integral membrane proteins by the following sequential process: in-solution acid hydrolysis, reverse phase chromatographic separation, trypsin or chymotrypsin digestion and nano-liquid chromatography–Fourier transform mass spectrometry. When we employed total membrane proteins of Synechocystis sp. PCC 6803, 155 integral membrane proteins out of a predictable 706 were identified in a single application, corresponding to 22% of a genome. The combined methods of acid hydrolysis-trypsin (AT) and acid hydrolysis-chymotrypsin (AC) identified both hydrophilic and hydrophobic domains of integral membrane proteins, respectively. The systematic approach revealed a more concrete data in mapping the repertoire of cyanobacterial membrane and membrane-linked proteome. [Copyright &y& Elsevier]

Abstract: The natural soil mineral conditioner illite effectively improved the germination and growth of lettuce when applied in particulate or powdered form. When illite was given in particulate and powdered forms, the germination rate of lettuce seeds improved remarkably up to 93% and 133%, respectively. Contrary to the developmental effects, the growth rate of lettuce treated with particulate illite improved slightly by 23%; powdered illite had no significant effects on lettuce growth rate. Thus, illite primarily affects seed germination rather than the growth of lettuce. To examine illite-induced proteins related to lettuce growth, differentially expressed proteins in lettuce leaves were analyzed by two-dimensional gel electrophoresis (2-DE) and matrix-assisted laser desorption ionization-time of flight/time of flight mass spectrometry (MALDI-TOF/TOF MS) followed by Mascot search. From the proteomic analysis, five down-regulated proteins were identified related to storage protein, carbon metabolism and energy conversion. Three up-regulated proteins were related to energy production/conversion and carbon fixation. These results demonstrate that illite treatment as a soil conditioner helps lettuce seed germination and lettuce growth by regulating carbon metabolic flux. [Copyright &y& Elsevier]

Abstract: Although recent advances in gel electrophoresis and mass spectrometry have greatly facilitated separation, purification, and identification of proteins, significant challenges remain in relation to phosphoprotein analysis. Here we introduce a powerful method for analysis of protein phosphorylation in which phosphorylation sites are labeled with guanidinoethanethiol (GET) by β-elimination/Michael addition prior to proteolysis and mass spectrometry (MS) analysis. This technique is especially useful in conjunction with gel-based technology in that all of the processes involved, including GET labeling, washing, and phosphospecific enzymatic hydrolysis, can be carried out in excised gel slices, thereby minimizing sample loss and contamination. The novel GET tag, which has a highly basic guanidine group, increases the peak intensities for the GET-labeled tryptic peptides by matrix-assisted laser desorption/ionization time-of-flight (MALDI–TOF) MS. In addition, phosphospecific proteolytic cleavage occurs at guanidinoethylcysteine (Gec) residue, which is arginine-mimic formed by GET tagging of phosphorylated serine residues. Thus, GET tagging is especially useful in analysis of long tryptic phosphopeptides. To illustrate the utility of the in-gel GET tagging and digestion approach, we used it to precisely analyze the phosphorylation sites of human glutathione S-transferase P1 (GSTP1), an enzyme involved in phase II metabolism of many carcinogens and anticancer drugs. The in-gel GET tagging/digestion technique significantly enhances the analytical potential of gel electrophoresis/MS in studies of proteome phosphorylation. [Copyright &y& Elsevier]

Alcoholic liver disease (ALD) and nonalcoholic fatty liver disease (NAFLD) are typically asymptomatic and slow-progressing but potentially fatal diseases that are common causes of liver cirrhosis and related complications. Exosomes are nano-sized extracellular vesicles that have been linked to various intercellular communication processes and can carry biological materials reflecting the state and severity of disease. In this study, shotgun proteomic analysis of the protein expression profiles of extracellular vesicles, including exosomes and microvesicles, enriched from human serum samples of 24 patients diagnosed with various fatty liver diseases was performed using liquid chromatography tandem mass spectrometry (LC-MS/MS) followed by protein identification and label-free quantification using the MaxQuant platform. A total of 329 proteins, including 190 previously reported exosome-specific proteins, were identified from four types of liver disease, where significant differences in protein expression were found in apolipoproteins, immunoglobulins, and other previously reported markers of liver disease. Principal component analysis of 61 proteins identified from MaxQuant analysis of the LC-MS/MS data provided a confident differentiation between ALD and NAFLD. The current investigation revealed the difference among various types of liver disease using LC-MS/MS of exosomes enriched from human serum samples of 24 patients where the most significantly up-regulation proteins were alpha-2-macroglobulin for alcoholic hepatitis and apolipoprotein C3 for nonalcoholic fatty liver disease. [Display omitted] • Exosomes were enriched from human serum samples with fatty liver diseases. • LC-MS/MS analyses of digested exosomal proteins were performed in triplicate. • A total of 329 proteins were identified from the enriched exosomes. • Confident differentiation was obtained between alcoholic liver disease and nonalcoholic fatty liver disease. [ABSTRACT FROM AUTHOR]

Citric acid (CA), as an organic chelator, plays a vital role in alleviating copper (Cu) stress-mediated oxidative damage, wherein a number of molecular mechanisms alter in plants. However, it remains largely unknown how CA regulates differentially abundant proteins (DAPs) in response to Cu stress in Brassica napus L. In the present study, we aimed to investigate the proteome changes in the leaves of B. L. seedlings in response to CA-mediated alleviation of Cu stress. Exposure of 21-day-old seedlings to Cu (25 and 50 μM) and CA (1.0 mM) for 7 days exhibited a dramatic inhibition of overall growth and considerable increase in the enzymatic activities (POD, SOD, CAT). Using a label-free proteome approach, a total of 6345 proteins were identified in differentially treated leaves, from which 426 proteins were differentially expressed among the treatment groups. Gene ontology (GO) and KEGG pathways analysis revealed that most of the differential abundance proteins were found to be involved in energy and carbohydrate metabolism, photosynthesis, protein metabolism, stress and defense, metal detoxification, and cell wall reorganization. Our results suggest that the downregulation of chlorophyll biosynthetic proteins involved in photosynthesis were consistent with reduced chlorophyll content. The increased abundance of proteins involved in stress and defense indicates that these DAPs might provide significant insights into the adaptation of Brassica seedlings to Cu stress. The abundances of key proteins were further verified by monitoring the mRNA expression level of the respective transcripts. Taken together, these findings provide a potential molecular mechanism towards Cu stress tolerance and open a new route in accelerating the phytoextraction of Cu through exogenous application of CA in B. napus. [ABSTRACT FROM AUTHOR]

Enhanced monitored natural attenuation was conducted with molasses injections to an in situ trichloroethylene (TCE)-contaminated groundwater site. Three pilot-scale wells were conducted for monitoring and substrate injections. At most 97.0% of the TCE was effectively degraded after about 600 days. TCE detected in the injection wells attained Tier 2 Groundwater Regulation in Taiwan for non-drinking water sources. The reaction rates of Cl–VOCs in one of the injection wells explained the mechanism how these dechlorinating species transferred TCE to cis-DCE and VC in anaerobic conditions. The magnitude of the 1st order kinetic growth rates of Dehalococcoides (Dhc) were consistent with the associated TCE degradability. The qPCR and T-RFLP results concluded existence of crucial dechlorinating species including Dhc, which performed critical level of 106 copy L−1 to trigger the TCE degradation. The detected t ceA and vcrA ranging from 2.07 × 103 to 6.57 × 105 copy L−1confirmed the pathway of TCE degradation. The two injection wells with the optimal TCE degradation showed increases of bacterial diversity estimated by the Shannon Index. The Shannon index explained the superiority of an increased bacterial diversity. The nonmetric multidimensional scaling results suggested that the bacterial dynamic was affected by the TCE degradation stages. The bacterial community structures were analogous when TCE was well degraded. • The molecular results provided evidence of existence of crucial dechlorinating species including Dehalococcoides spp. • Dehalococcoides spp. With the critical abundance of 106 copy L−1 was critical for TCE degradation. • Detections of tceA (as high as 107 copy L−1) and vcrA confirmed the pathway of TCE degradation. • The optimal TCE degradation was observed with increases of bacterial diversity estimated by the Shannon Index. • Nonmetric multidimensional scaling result indicated the bacterial community dynamic depended on the TCE degradation stages. [ABSTRACT FROM AUTHOR]