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Protein electrophoresis

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https://www.readbyqxmd.com/read/29777872/inventory-of-proteoforms-as-a-current-challenge-of-proteomics-some-technical-aspects
#1
Stanislav Naryzhny
The main intricacy in the human proteome is that it is tremendously complex and composed from diverse and heterogeneous gene products. These products are called protein species or proteoforms and are the smallest units of the proteome. In pursuit of the comprehensive profiling of the human proteome, significant advances in the technology of so called "Top-Down" mass spectrometry based proteomics, have been made. However, the scale of performance of this approach is still far behind the "Bottom-Up", peptide-centric techniques...
May 16, 2018: Journal of Proteomics
https://www.readbyqxmd.com/read/29776310/preparation-and-evaluation-of-ribonuclease-resistant-viral-hiv-rna-standards-based-on-armored-rna-technology
#2
Mohammad Gholami, Mehrdad Ravanshad, Kazem Baesi, Siamak M. Samiee, Negin Hosseini Rozbahani, Minoo Mohraz
BAckground: The human immunodeficiency virus type 1 (HIV-1) is an infectious viral agent that gradually extinguishes the immune system, resulting in acquired immune deficiency syndrome (AIDS). The aim of this study was to construct an RNA-positive control based on armored (AR) RNA technology, using HIV-1 RNA as a model. Methods: The MS2 maturase, a coat protein gene (at positions 1765 to 1787) and HIV-1 pol gene were cloned into pET-32a plasmid. The prepared plasmid was transformed into Escherichia coli strain BL2 (DE3), and the expression of the construct was induced by 1 mM of isopropyl-L-thio-D-galactopyranoside (IPTG) at 37 °C for 16 h to obtain the fabricated AR RNA...
May 19, 2018: Iranian Biomedical Journal
https://www.readbyqxmd.com/read/29775569/modification-of-chickpea-cystatin-by-reactive-dicarbonyl-species-glycation-oxidation-and-aggregation
#3
Sheraz Ahmad Bhat, Waseem Feeroze Bhat, Mohammad Afsar, Mohd Shahnawaz Khan, Bilqees Bano
Reactive dicarbonyl species such as methylglyoxal (MGO) and glyoxal (GO) have recently received extensive attention due to their high reactivity and ability to modify biological substances such as proteins, phospholipids, and DNA. In case of proteins these reactive species mainly react with lysine and arginine residues to form AGEs, oxidative products, and aggregates. Chickpea cystatin (CPC) was incubated with varying concentrations of glyoxal and methylglyoxal which caused, along with altered secondary and tertiary structures, glycation, functional inactivation, altered redox state, cross-linking and high-molecular-mass aggregation...
May 15, 2018: Archives of Biochemistry and Biophysics
https://www.readbyqxmd.com/read/29775289/mass-spectral-detection-of-diethoxyphospho-tyrosine-adducts-on-proteins-from-hek293-cells-using-monoclonal-antibody-depy-for-enrichment
#4
Seda Onder, Lawrence M Schopfer, Ozden Tacal, Thomas A Blake, Rudolph C Johnson, Oksana Lockridge
Chronic illness from exposure to organophosphorus toxicants is hypothesized to involve modification of unknown proteins. Tyrosine in proteins that have no active site serine readily reacts with organophosphorus toxicants. We developed a monoclonal antibody, depY, that specifically recognizes diethoxyphospho-tyrosine in proteins and peptides, independent of the surrounding amino acid sequence. Our goal in the current study was to identify diethoxyphosphorylated proteins in human HEK293 cell lysate treated with chlorpyrifos oxon...
May 18, 2018: Chemical Research in Toxicology
https://www.readbyqxmd.com/read/29774828/bilberry-pomace-in-rabbit-nutrition-effects-on-growth-performance-apparent-digestibility-caecal-traits-bacterial-community-and-antioxidant-status
#5
S Dabbou, I Ferrocino, A Kovitvadhi, S Bergagna, D Dezzuto, A Schiavone, L Cocolin, F Gai, V Santoro, L Gasco
Agricultural by-products could be used as alternative raw materials in rabbit nutrition as they have been found to be highly nutritious and low cost feeding sources. The aim of this study was to estimate the nutritive value and potential use of bilberry pomace (BP) for growing rabbits. A total of 144 Grimaud rabbits (35 days old) were allotted to four groups and fed with a diet containing increasing level of BP: BP0 (basal diet), BP5, BP10 and BP15 containing 0, 50, 100 and 150 g/kg respectively. Growth trial lasted 48 days; apparent digestibility was evaluated, starting at 46 days of age, over 4 consecutive days...
May 18, 2018: Animal: An International Journal of Animal Bioscience
https://www.readbyqxmd.com/read/29774437/lexa-binds-to-transcription-regulatory-site-of-cell-division-gene-ftsz-in-toxic-cyanobacterium-microcystis-aeruginosa
#6
Takashi Honda, Daichi Morimoto, Yoshihiko Sako, Takashi Yoshida
Previously, we showed that DNA replication and cell division in toxic cyanobacterium Microcystis aeruginosa are coordinated by transcriptional regulation of cell division gene ftsZ and that an unknown protein specifically bound upstream of ftsZ (BpFz; DNA-binding protein to an upstream site of ftsZ) during successful DNA replication and cell division. Here, we purified BpFz from M. aeruginosa strain NIES-298 using DNA-affinity chromatography and gel-slicing combined with gel electrophoresis mobility shift assay (EMSA)...
May 17, 2018: Marine Biotechnology
https://www.readbyqxmd.com/read/29773679/comparative-proteomic-analysis-reveals-the-regulatory-network-of-the-vea-gene-during-asexual-and-sexual-spore-development-of-aspergillus-cristatus
#7
Hui Liu, Shilei Sang, Hui Wang, Xiyi Ren, Yumei Tan, Wei Chen, Zuoyi Liu, Yongxiang Liu
Aspergillus cristatus belongs to the homothallic fungal group that undergoes a sexual stage without asexual conidiation under hypotonic conditions, while hypertonic medium induces initiation of the asexual stage and completely blocks sexual development. However, the veA deletion mutant only produces conidia in hypotonic medium after a 24 h culture, but both asexual and sexual spores are observed after 72 h. To elucidate the molecular mechanism of how VeA regulates asexual and sexual spore development in A. cristatus , two dimensional electrophoresis combined with matrix-assisted laser desorption/ionization-tandem time of flight mass spectrometry analysis were applied to identify 173 differentially expressed proteins (DEPs) by comparing the agamotype (24 h) and teleomorph (72 h) with wild-type A...
May 17, 2018: Bioscience Reports
https://www.readbyqxmd.com/read/29772786/tau-fibril-formation-in-cultured-cells-compatible-with-a-mouse-model-of-tauopathy
#8
Gen Matsumoto, Kazuki Matsumoto, Taeko Kimura, Tetsuya Suhara, Makoto Higuchi, Naruhiko Sahara, Nozomu Mori
Neurofibrillary tangles composed of hyperphosphorylated tau protein are primarily neuropathological features of a number of neurodegenerative diseases collectively termed tauopathy. To understand the mechanisms underlying the cause of tauopathy, precise cellular and animal models are required. Recent data suggest that the transient introduction of exogenous tau can accelerate the development of tauopathy in the brains of non-transgenic and transgenic mice expressing wild-type human tau. However, the transmission mechanism leading to tauopathy is not fully understood...
May 17, 2018: International Journal of Molecular Sciences
https://www.readbyqxmd.com/read/29772469/synthesis-of-steroidal-imidazolidinthiones-as-potential-apoptotic-agents-investigation-by-theoretical-and-experimental-studies
#9
Ayaz Mahmood Dar, Rizwan Nabi, Shafia Mir, Manzoor Ahmad Gatoo, Shamsuzzaman, Shabir H Lone
New steroidal imidazolidinthione derivatives (4-6) were synthesized from steroidal thiosemicarbazones and dichloroethane. The synthesized compounds were characterized using spectral data analysis. Theoretical DFT involving B3LYP/6-31G∗∗ level of theory was employed to gain insights into the molecular structure of the target compounds. MEPS and FMO analysis were carried out. HOMO-LUMO energy gap was determined which helped to evaluate various global descriptors like hardness, chemical potential, electronegativity, nucleophilicity and electrophilicity index, etc...
May 2, 2018: Bioorganic Chemistry
https://www.readbyqxmd.com/read/29771309/identification-of-a-specific-domain-of-porphyromonas-gingivalis-hgp44-responsible-for-adhesion-to-treponema-denticola
#10
Kouki Yoshikawa, Yuichiro Kikuchi, Eitoyo Kokubu, Kentaro Imamura, Atsushi Saito, Kazuyuki Ishihara
Interaction between two periodontal pathogens, Porphyromonas gingivalis and Treponema denticola, contributes to plaque biofilm formation. P. gingivalis forms aggregates with T. denticola through its adhesion/hemagglutinin domain (Hgp44). In this study, we investigated the specific domain of P. gingivalis Hgp44 responsible for adhesion to T. denticola using expression vectors harboring P. gingivalis Hgp44 DNA sequences encoding amino acid residues 1-419. Six plasmids harboring fragments in this region were generated by PCR amplification and self-ligation, and recombinant proteins r-Hgp44 (residues 1-419), r-Hgp441 (residues 1-124), r-Hgp442 (1-199), r-Hgp443 (1-316), r-Hgp444 (199-419), r-Hgp445 (124-198), and r-Hgp446 (199-316) were produced, as confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting...
May 16, 2018: Pathogens and Disease
https://www.readbyqxmd.com/read/29771257/characterization-of-mercury-binding-proteins-in-rat-blood-plasma
#11
Yiling Li, Bin He, Qiying Nong, Guangbo Qu, Lihong Liu, Jianbo Shi, Ligang Hu, Guibin Jiang
Mercury-binding protein profiles in rat plasma at different levels of mercury exposure in vitro and in vivo were systematically investigated using column gel electrophoresis coupled with inductively coupled plasma-mass spectrometry. The current finding provided various protein candidates that may play critical roles in mercury binding and transporting in plasma.
May 16, 2018: Chemical Communications: Chem Comm
https://www.readbyqxmd.com/read/29770965/determination-of-protein-charge-variants-with-imaged-capillary-isoelectric-focusing-and-capillary-zone-electrophoresis
#12
REVIEW
Julia Kahle, Hermann Wätzig
Biopharmaceuticals are increasingly present in pharmaceutical therapy. These highly complex molecules are still challenging for development and quality control. Charge heterogeneity is next to size heterogeneity, glycan analysis or peptide mapping one main aspect of their thorough characterization. This review focuses on the charge-based analysis of biopharmaceuticals through capillary electrophoresis. An overview is given about the three separation techniques cIEF, icIEF and CZE for the analyses of charge variants, emphasizing assays and their precision...
May 17, 2018: Electrophoresis
https://www.readbyqxmd.com/read/29767655/proteomic-analysis-of-stachyose-contribution-to-the-growth-of-lactobacillus-acidophilus-cicc22162
#13
Xian-Feng Zhong, Yu-Bo Zhang, Gui-Dong Huang, Yong-Zhong Ouyang, Dong-Jiang Liao, Jia-Wei Peng, Wei-Zhi Huang
Stachyose is a functional oligosaccharide, acting as a potential prebiotic for colonic fermentation. To understand the mechanism of how stachyose promotes the growth of probiotic bacterium, we analyzed the differences of the proteome of Lactobacillus acidophilus grown on stachyose or glucose. By a combination of two-dimensional electrophoresis and mass spectrometry analysis, we observed 16 proteins differentially abundant under these two conditions and identified 9 protein spots. Six of these proteins were highly abundant when stachyose was used as the sole carbon source...
May 16, 2018: Food & Function
https://www.readbyqxmd.com/read/29766032/serum-free-erythroid-differentiation-for-efficient-genetic-modification-and-high-level-adult-hemoglobin-production
#14
Naoya Uchida, Selami Demirci, Juan J Haro-Mora, Atsushi Fujita, Lydia N Raines, Matthew M Hsieh, John F Tisdale
In vitro erythroid differentiation from primary human cells is valuable to develop genetic strategies for hemoglobin disorders. However, current erythroid differentiation methods are encumbered by modest transduction rates and high baseline fetal hemoglobin production. In this study, we sought to improve both genetic modification and hemoglobin production among human erythroid cells in vitro . To model therapeutic strategies, we transduced human CD34+ cells and peripheral blood mononuclear cells (PBMCs) with lentiviral vectors and compared erythropoietin-based erythroid differentiation using fetal-bovine-serum-containing media and serum-free media...
June 15, 2018: Molecular Therapy. Methods & Clinical Development
https://www.readbyqxmd.com/read/29763592/coomassie-does-it-better-a-robin-hood-approach-to-total-protein-quantification
#15
Nour Noaman, Jens R Coorssen
Quantitative comparative proteomics require accurate and reproducible assessments of total protein concentration. We report a straightforward, cost-effective adaptation of an established commercial method for total protein quantification, utilising non-proprietary materials and colloidal Coomassie Brilliant Blue (cCBB) staining to achieve greater reproducibility, equal sensitivity, and optimal linearity of signal within a practical concentration range for proteins in common solubilisation buffers (i.e. for isoelectric focussing and/or SDS-PAGE)...
May 12, 2018: Analytical Biochemistry
https://www.readbyqxmd.com/read/29761223/serum-profile-of-transferrin-isoforms-in-juvenile-idiopathic-arthritis-a-preliminary-study
#16
Ewa Gruszewska, Magdalena Sienkiewicz, Paweł Abramowicz, Jerzy Konstantynowicz, Monika Gudowska-Sawczuk, Lech Chrostek, Bogdan Cylwik
It is reported that alterations in protein glycosylation are present in adult rheumatic diseases; however, the data related to pediatric rheumatic conditions are very scarce. The aim of this study was to assess the effect of juvenile idiopathic arthritis (JIA) on the serum glycosylation profile of transferrin isoforms. Twenty-five patients with different clinical forms of an active JIA and 22 healthy controls were studied. Serum samples were analyzed by capillary electrophoresis on MINICAP electrophoretic system (Sebia, France) to determine the levels of transferrin isoforms...
May 14, 2018: Rheumatology International
https://www.readbyqxmd.com/read/29759138/impact-of-imipramine-on-proteome-of-rat-primary-glial-cells
#17
Sylwia Kedracka-Krok, Bianka Swiderska, Anna M Bielecka-Wajdman, Gabriela Prus, Bozena Skupien-Rabian, Urszula Jankowska, Ewa Obuchowicz
Microglia and astrocytes, two types of glial cells are known to be important targets for antidepressant drugs. Here we used a comprehensive proteomic analysis to examine the effect of imipramine on rat primary mixed glial culture. The two-dimensional differential gel electrophoresis method allowed us to identify 62 proteins that were altered by imipramine. Functional analysis revealed that imipramine influenced the level of proteins involved in oxidative stress; in particular, it elevated the level of glutathione transferases...
July 15, 2018: Journal of Neuroimmunology
https://www.readbyqxmd.com/read/29758474/purification-and-characterization-of-two-pathogenesis-related-class-10-protein-isoforms-with-ribonuclease-activity-from-the-fresh-angelica-sinensis-roots
#18
Jianru Pan, Xiangling Wang, Lingling Li, Xian Li, Xiaoqiang Ye, Di Lv, Cuihuang Chen, Shutao Liu, Huocong He
In this study, two pathogenesis-related (PR) class 10 protein isoforms, ASPR-1 and ASPR-2, were purified from fresh roots of the Chinese medicinal plant Angelica sinensis (A. sinensis) using 80% ammonium sulfate precipitation, Sephadex G50 gel filtration chromatography, and DEAE-Sepharose ion-exchange chromatography. The molecular masses of ASPR-1 and ASPR-2 were estimated to be 16.66 kDa and 16.46 kDa, respectively, using sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The isoforms are both glycoproteins containing glycosyl contents of 1...
April 30, 2018: Plant Physiology and Biochemistry: PPB
https://www.readbyqxmd.com/read/29757586/production-of-recombinant-protein-of-salsola-kali-sal-k1-pollen-allergen-in-lactococcus-lactis
#19
Ziba Ghasemi, Abdol-Reza Varasteh, Malihe Moghadam, Farnaz Sedghi, Farahzad Jabbariazad, Mojtaba Sankian
The Salsola kali pollen is considered the main cause of allergic sensitization in desert and semi-desert regions. We have constructed recombinant Lactococcus lactis producing Sal k1 protein with the aim of using it as a mucosal vaccine for specific immunotherapy. The Sal k1 gene was amplified, and transferred into a PNZ 8148 plasmid. The PNZ8148-Sal k1 recombinant plasmid was transformed into competent E.coli strain MC1061 for replication, and then was isolated and cloned into competent L. lactis by electroporation...
April 2018: Iranian Journal of Allergy, Asthma, and Immunology
https://www.readbyqxmd.com/read/29757289/use-of-two-dimensional-semi-denaturing-detergent-agarose-gel-electrophoresis-to-confirm-size-heterogeneity-of-amyloid-or-amyloid-like-fibers
#20
Sarah Hanna-Addams, Zhigao Wang
Amyloid or amyloid-like fibers have been associated with many human diseases, and are now being discovered to be important for many signaling pathways. The ability to readily detect the formation of these fibers under various experimental conditions is essential for understanding their potential function. Many methods have been used to detect the fibers, but not without some drawbacks. For example, electron microscopy (EM), or staining with Congo Red or Thioflavin T often requires purification of the fibers...
April 26, 2018: Journal of Visualized Experiments: JoVE
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