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https://www.readbyqxmd.com/read/28735093/stable-isotope-labeling-by-fatty-acids-in-cell-culture-silfac-coupled-with-isotope-pattern-dependent-mass-spectrometry-for-global-screening-of-lipid-hydroperoxide-mediated-protein-modifications
#1
Ryo Takahashi, Shuhei Fujioka, Tomoyuki Oe, Seon Hwa Lee
Lipid hydroperoxide-mediated modifications of proteins are receiving increasing attention because of their possible involvement in various degenerative diseases. These biological effects are attributed to the ability of lipid peroxidation-derived aldehydes to react with the nucleophilic sites of proteins. Here we describe a methodology involving metabolic labeling coupled with mass spectrometry-based proteomic analysis that enables global screening of lipid hydroperoxide-mediated protein modifications in a cell system...
July 19, 2017: Journal of Proteomics
https://www.readbyqxmd.com/read/28735080/recombinant-truncated-e-protein-as-a-new-vaccine-candidate-against-nontypeable-h-influenzae-its-expression-and-immunogenic-evaluation
#2
Ava Behrouzi, Saeid Bouzari, Farzam Vaziri, Abolfazl Fateh, Parviz Afrough, Atefeh Davoudi Vijeh Motlagh, Seyed Davar Siadat
Protein E (PE) is a conserved entity observed in both nontypeable Haemophilus influenzae (NTHi) and encapsulated H. influenzae. This is a small surface lipoprotein, consisting of only 160 amino acids, involved in the adhesion of H. influenzae to various types of epithelial cells. A 384-bp-long fragment from NTHi PE was cloned into the prokaryotic expression vector pBAD-gIIIA. The recombinant protein was expressed with arabinose and then purified by affinity purification on an Ni-NTA agarose matrix. BALB/c mice were immunized by subcutaneous injection with purified recombinant truncated PE mixed with an alum adjuvant...
July 19, 2017: Microbial Pathogenesis
https://www.readbyqxmd.com/read/28734840/an-improved-purification-method-for-the-lysosomal-storage-disease-protein-%C3%AE-glucuronidase-produced-in-cho-cells
#3
Erica J Fratz-Berilla, Stephanie A Ketcham, Hamideh Parhiz, Muhammad Ashraf, Chikkathur N Madhavarao
Human β-glucuronidase (GUS; EC 3.2.1.31) is a lysosomal enzyme that catalyzes the hydrolysis of β-d-glucuronic acid residues from the non-reducing termini of glycosaminoglycans. Impairment in GUS function leads to the metabolic disorder mucopolysaccharidosis type VII, also known as Sly syndrome. We produced GUS from a CHO cell line grown in suspension in a 15 L perfused bioreactor and developed a three step purification procedure that yields ∼99% pure enzyme with a recovery of more than 40%. The method can be completed in two days and has the potential to be integrated into a continuous manufacturing scheme...
July 19, 2017: Protein Expression and Purification
https://www.readbyqxmd.com/read/28734838/expression-purification-and-characterization-of-sll1981-protein-from-cyanobacterium-synechocystis-sp-pcc6803
#4
Xiaoqin Wang, Guofeng Lei, Xiaoyu Wu, Fei Wang, Chongde Lai, Zhimin Li
The sll1981 protein from cyanobacterium Synechocystis sp. PCC6803 had been reported to exhibit acetolactate synthase (ALS) and L-myo-inositol-1-phosphate synthase (MIPS) activities previously. Based on amino acids sequences alignment, sll1981 protein was postulated to function as α-ketoglutarate decarboxylase (α-KGD), which played important role in completing cyanobacterial tricarboxylic acid (TCA) cycle. However the detailed enzymatic kinetics of sll1981 as ALS, MIPS and α-KGD were not determined yet. In this study, the recombinant sll1981 protein was purified from supernatant of E...
July 19, 2017: Protein Expression and Purification
https://www.readbyqxmd.com/read/28734361/preparation-of-intact-mitochondria-using-free-flow-isoelectric-focusing-with-post-ph-gradient-sample-injection-for-morphological-functional-and-proteomics-studies
#5
Yu-Chen He, Fan-Zhi Kong, Liu-Yin Fan, Jane Y Wu, Xiao-Ping Liu, Jun Li, Yan Sun, Qiang Zhang, Ying Yang, Xue-Jing Wu, Hua Xiao, Cheng-Xi Cao
Mitochondria play essential roles in both energy metabolism and cell signaling, which are critical for cell survival. Although significant efforts have been invested in understanding mitochondrial biology, methods for intact mitochondria preparation are technically challenging and remain to be improved. New methods for heterogeneous mitochondria purification will therefore boost our understanding on their physiological and biophysical properties. Herein, we developed a novel recycling free-flow isoelectric focusing (RFFIEF) with post-pH gradient sample injection (post-PGSI) for preparative separation of mitochondria...
August 22, 2017: Analytica Chimica Acta
https://www.readbyqxmd.com/read/28734049/development-of-aptamers-against-unpurified-proteins
#6
Shinichi Goto, Kaori Tsukakoshi, Kazunori Ikebukuro
SELEX (Systematic Evolution of Ligands by EXponential enrichment) has been widely used for the generation of aptamers against target proteins. However, its requirement for pure target proteins remains a major problem in aptamer selection, as procedures for protein purification from crude bio-samples are not only complicated but also time and labour consuming. This is because native proteins can be found in a large number of diverse forms because of posttranslational modifications and their complicated molecular conformations...
July 22, 2017: Biotechnology and Bioengineering
https://www.readbyqxmd.com/read/28733633/high-fidelity-de-novo-synthesis-of-pathways-using-microchip-synthesized-oligonucleotides-and-general-molecular-biology-equipment
#7
Wen Wan, Min Lu, Dongmei Wang, Xiaolian Gao, Jiong Hong
Engineering and evaluation of synthetic routes for generating valuable compounds require accurate and cost-effective de novo synthesis of genetic pathways. Here, we present an economical and streamlined de novo DNA synthesis approach for engineering a synthetic pathway with microchip-synthesized oligonucleotides (oligo). The process integrates entire oligo pool amplification, error-removal, and assembly of long DNA molecules. We utilized this method to construct a functional lycopene biosynthetic pathway (11...
July 21, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28733289/an-enterotoxin-like-binary-protein-from-pseudomonas-protegens-with-potent-nematicidal-activity
#8
Jun-Zhi Wei, Daniel L Siehl, Zhenglin Hou, Barbara Rosen, Jarred Oral, Christopher G Taylor, Gusui Wu
Soil microbes are a major food source for free-living soil nematodes. It is known that certain soil bacteria have evolved systems to combat predation. We identified the nematode-antagonistic Pseudomonas protegens strain 15G2 from screening of microbes. Through protein purification we identified a binary protein, designated as Pp-ANP, that is responsible for the nematicidal activity. This binary protein inhibits Caenorhabditis elegans growth and development by arresting larvae at L1 stage and killing older staged worms...
July 21, 2017: Applied and Environmental Microbiology
https://www.readbyqxmd.com/read/28731846/presence-and-future-of-culturing-bacteria
#9
Jörg Overmann, Birte Abt, Johannes Sikorski
The cultivation of bacteria is highly biased toward a few phylogenetic groups. Many of the currently underexplored bacterial lineages likely have novel biosynthetic pathways and unknown biochemical features. New cultivation concepts have been developed based on an improved understanding of the ecology of previously not-cultured bacteria. Particularly successful were improved media that mimic the natural types and concentrations of substrates and nutrients, high-throughput cultivation techniques, and approaches that exploit biofilm formation and bacterial interactions...
July 21, 2017: Annual Review of Microbiology
https://www.readbyqxmd.com/read/28731574/rapid-buffer-and-ligand-screening-for-affinity-chromatography-by-multiplexed-surface-plasmon-resonance-imaging
#10
Karin P M Geuijen, Daniëlle E J W van Wijk-Basten, David F Egging, Richard B M Schasfoort, Michel H Eppink
Protein purifications are often based on the principle of affinity chromatography, where the protein of interest selectively binds to an immobilized ligand. The development of affinity purification requires selecting proper wash and elution conditions. In recent years, miniaturization of the purification process is applied to speed up the development (e.g. microtiterplates, robocolumns). We have studied the application of surface plasmon resonance imaging (SPRi) as a tool to simultaneously screen many buffer conditions for wash and elution steps in an affinity-based purification process...
July 21, 2017: Biotechnology Journal
https://www.readbyqxmd.com/read/28731571/model-based-investigation-on-the-mass-transfer-and-adsorption-mechanisms-of-mono-pegylated-lysozyme-in-ion-exchange-chromatography
#11
Josefine Morgenstern, Gang Wang, Pascal Baumann, Jürgen Hubbuch
Recent studies highlighted the potential of PEGylated proteins to improve stabilities and pharmacokinetics of protein drugs. Ion-exchange chromatography (IEX) is among the most frequently used purification methods for PEGylated proteins. However, the underlying physical mechanisms allowing for a separation of different PEGamers (proteins with a varying number of attached PEG molecules) are not yet fully understood. In this work, mechanistic chromatography modeling was applied to gain a deeper understanding of the mass transfer and adsorption/desorption mechanisms of mono-PEGylated proteins in IEX...
July 21, 2017: Biotechnology Journal
https://www.readbyqxmd.com/read/28731485/th17-regulatory-t-cell-imbalance-in-sepsis-patients-with-multiple-organ-dysfunction-syndrome-attenuated-by-high-volume-hemofiltration
#12
Jiguang Guo, Wu Tao, Dan Tang, Jianbin Zhang
BACKGROUND: We assessed the Th17 (T-helper cell)/Treg (Regulatory T cell) imbalance in sepsis patients with multiple organ dysfunction syndrome (MODS) and the clinical benefits of continuous high-volume hemofiltration (HVHF). METHODS: 48 sepsis patients, including 22 patients with MODS (MODS group, n = 22) and 26 without (non-MODS group, n = 26), and 20 healthy volunteer controls were enrolled. The patients in MODS group were randomly divided into the continuous blood purification (CBP) group (n = 11) receiving conventional comprehensive treatment plus high-volume hemofiltration and the non-CBP group (n = 11) receiving conventional comprehensive treatment only...
July 14, 2017: International Journal of Artificial Organs
https://www.readbyqxmd.com/read/28731187/isolation-and-purification-of-novel-peptides-derived-from-sepia-ink-effects-on-apoptosis-of-prostate-cancer-cell-pc%C3%A2-3
#13
Fangfang Huang, Yinwen Jing, Guofang Ding, Zuisu Yang
Novel prostate cancer therapeutics are in high demand. In order to identify potential therapeutic targets, protein from sepia ink was hydrolyzed by utilizing pepsin in an orthogonal array design. Pepsin hydrolysate (SH) obtained at optimal conditions exhibited the highest antitumor activity. Subsequently, a novel antitumor peptide, which was termed SHP, was isolated through ultrafiltration, gel filtration chromatography and reversed phase high‑performance liquid chromatography. The amino acid sequence of SHP was identified as Leu‑Lys‑Glu‑Glu‑Asn‑Arg‑Arg‑Arg‑Arg‑Asp with a molecular mass of 1371...
July 21, 2017: Molecular Medicine Reports
https://www.readbyqxmd.com/read/28730817/efficient-solid-state-electrochemiluminescence-from-high-quality-perovskite-quantum-dot-films
#14
Jingjing Xue, Ziyi Zhang, Fenfen Zheng, Qin Xu, Jinchun Xu, Guizheng Zou, Lingling Li, Jun-Jie Zhu
Halide perovskite materials have emerged as a new class of revolutionary photovoltaic and optoelectronic nanomaterials. However, the study on electrochemiluminescence (ECL) from halide perovskite nanomaterials is still in its infancy due to their instability, sensitivity, and difficulties in purification and film formation. Here, we propose a scraping coating method for the fabrication of high-quality halide perovskite quantum dot (QD) film on electrode, which shows dense and uniform packing with minimum grain size...
July 24, 2017: Analytical Chemistry
https://www.readbyqxmd.com/read/28730714/alkaloid-profiling-of-herbal-drugs-using-high-resolution-mass-spectrometry
#15
Tiziana Nardin, Edi Piasentier, Chiara Barnaba, Roberto Larcher
Herbal infusions are consumed worldwide thanks to their "natural" beneficial effects, also due to the presence of alkaloids, although these compounds can have poisonous effects. A method combining online solid-phase purification with high resolution mass spectrometry was used to define the alkaloid profiles of 117 herbs and 7 commercial blends. 41 alkaloids were quantified in reference to analytical standards, while the presence of a further 116 was confirmed on the basis of accurate mass, retention time and fragmentation profile...
July 21, 2017: Drug Testing and Analysis
https://www.readbyqxmd.com/read/28730361/the-effect-of-different-surface-materials-on-runoff-quality-in-permeable-pavement-systems
#16
Haiyan Li, Zhifei Li, Xiaoran Zhang, Zhuorong Li, Dongqing Liu, Tanghu Li, Ziyang Zhang
To investigate the effect of different permeable pavement surface materials on the removal of pollutants from urban storm-runoff, six commonly surface materials (porous asphalt, porous concrete, cement brick, ceramic brick, sand base brick, and shale brick) were selected in this study and the research was carried out by column experiments. Except the concentrations of total suspended solids (TSS), chemical oxygen demand (COD), ammonia nitrogen (NH4-N), nitrate nitrogen (NO3-N), total nitrogen (TN), and total phosphorus (TP) in the influent and effluent that were measured, the removal mechanism of pollutants was discussed further...
July 20, 2017: Environmental Science and Pollution Research International
https://www.readbyqxmd.com/read/28730304/novel-ribonuclease-activity-of-cusativin-from-cucumis-sativus-for-mapping-nucleoside-modifications-in-rna
#17
Balasubrahmanyam Addepalli, Sarah Venus, Priti Thakur, Patrick A Limbach
A recombinant ribonuclease, cusativin, was characterized for its cytidine-specific cleavage ability of RNA to map chemical modifications. Following purification of native cusativin protein as described before (Rojo et al. Planta 194:328, 17), partial amino acid sequencing was carried out to identify the corresponding protein coding gene in cucumber genome. Cloning and heterologous expression of the identified gene in Escherichia coli resulted in successful production of active protein as a C-terminal His-tag fusion protein...
July 20, 2017: Analytical and Bioanalytical Chemistry
https://www.readbyqxmd.com/read/28730213/magnetic-separation-its-application-in-mining-waste-purification-medicine-biochemistry-and-chemistry
#18
REVIEW
M Iranmanesh, J Hulliger
The use of strong magnetic field gradients and high magnetic fields generated by permanent magnets or superconducting coils has found applications in many fields such as mining, solid state chemistry, biochemistry and medical research. Lab scale or industrial implementations involve separation of macro- and nanoparticles, cells, proteins, and macromolecules down to small molecules and ions. Most promising are those attempts where the object to be separated is attached to a strong magnetic nanoparticle. Here, all kinds of specific affinity interactions are used to attach magnetic carrier particles to mainly objects of biological interest...
July 21, 2017: Chemical Society Reviews
https://www.readbyqxmd.com/read/28729719/inhibition-of-tgf-%C3%AE-signaling-supports-high-proliferative-potential-of-diverse-p63-mouse-epithelial-progenitor-cells-in-vitro
#19
Daisuke Suzuki, Filipa Pinto, Makoto Senoo
Mouse models have been used to provide primary cells to study physiology and pathogenesis of epithelia. However, highly efficient simple approaches to propagate mouse primary epithelial cells remain challenging. Here, we show that pharmacological inhibition of TGF-β signaling enables long-term expansion of p63(+) epithelial progenitor cells in low Ca(2+) media without the need of progenitor cell-purification steps or support by a feeder cell layer. We find that TGF-β signaling is operative in mouse primary keratinocytes in conventional cultures as determined by the nuclear Smad2/3 localization...
July 20, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28729020/purification-of-cell-culture-derived-influenza-a-virus-via-continuous-anion-exchange-chromatography-on-monoliths
#20
Laura M Fischer, Michael W Wolff, Udo Reichl
The continuously increasing demand for potent and safe vaccines and the intensifying economic pressure on health care systems underlines the need for further optimization of vaccine manufacturing. Here, we focus on downstream processing of human influenza vaccines, investigating the purification of serum-free cell culture-derived influenza virus (A/PR/8/34 H1N1) using continuous chromatography. Therefore, quaternary amine anion exchange monoliths (CIM® QA) were characterized for their capacity to capture virus particles from animal cells cultivated in different media and their ability to separate virions from contaminating host cell proteins and DNA...
July 17, 2017: Vaccine
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