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https://www.readbyqxmd.com/read/29142197/proteotoxicity-in-cardiac-amyloidosis-amyloidogenic-light-chains-affect-the-levels-of-intracellular-proteins-in-human-heart-cells
#1
Esther Imperlini, Massimiliano Gnecchi, Paola Rognoni, Eduard Sabidò, Maria Chiara Ciuffreda, Giovanni Palladini, Guadalupe Espadas, Francesco Mattia Mancuso, Margherita Bozzola, Giuseppe Malpasso, Veronica Valentini, Giuseppina Palladini, Stefania Orrù, Giovanni Ferraro, Paolo Milani, Stefano Perlini, Francesco Salvatore, Giampaolo Merlini, Francesca Lavatelli
AL amyloidosis is characterized by widespread deposition of immunoglobulin light chains (LCs) as amyloid fibrils. Cardiac involvement is frequent and leads to life-threatening cardiomyopathy. Besides the tissue alteration caused by fibrils, clinical and experimental evidence indicates that cardiac damage is also caused by proteotoxicity of prefibrillar amyloidogenic species. As in other amyloidoses, the damage mechanisms at cellular level are complex and largely undefined. We have characterized the molecular changes in primary human cardiac fibroblasts (hCFs) exposed in vitro to soluble amyloidogenic cardiotoxic LCs from AL cardiomyopathy patients...
November 15, 2017: Scientific Reports
https://www.readbyqxmd.com/read/29131589/using-tryptophan-mutants-to-probe-the-structural-and-functional-status-of-bssco-a-copper-binding-cytochrome-c-oxidase-assembly-protein-from-bacillus-subtilis
#2
Shina Hussain, Diann Andrews, Bruce Charles Hill
The Synthesis of Cytochrome Oxidase protein from Bacillus subtilis (i.e., BsSCO) binds copper with pM affinity, which increases the protein's melting temperature (i.e., TM) by 20 oC. Here two native tryptophans (i.e., W36 and W101) are identified as major contributors to BsSCO's structural form and their contributions to stability, intrinsic fluorescence and copper binding properties of BsSCO are explored. Single mutations of tryptophan to phenylalanine decrease TM by 10 oC and folding free energy by 3-4 kcal/mol...
November 13, 2017: Biochemistry
https://www.readbyqxmd.com/read/29121649/spatiotemporal-dynamics-of-hsv-genome-nuclear-entry-and-compaction-state-transitions-using-bioorthogonal-chemistry-and-super-resolution-microscopy
#3
Eiki Sekine, Nora Schmidt, David Gaboriau, Peter O'Hare
We investigated the spatiotemporal dynamics of HSV genome transport during the initiation of infection using viruses containing bioorthogonal traceable precursors incorporated into their genomes (HSVEdC). In vitro assays revealed a structural alteration in the capsid induced upon HSVEdC binding to solid supports that allowed coupling to external capture agents and demonstrated that the vast majority of individual virions contained bioorthogonally-tagged genomes. Using HSVEdC in vivo we reveal novel aspects of the kinetics, localisation, mechanistic entry requirements and morphological transitions of infecting genomes...
November 9, 2017: PLoS Pathogens
https://www.readbyqxmd.com/read/29121483/effects-of-n-carbamylglutamate-and-l-arginine-on-gonadotrophin-releasing-hormone-gnrh-gene-expression-and-secretion-in-gt1-7-cells
#4
Y Liu, J H Bai, X L Xu, Z L Chen, L J Spicer, T Feng
Recent studies have shown that N-carbamylglutamate (NCG) and arginine (ARG) supplementation improves reproductive performance in livestock. The objectives of the present study were to evaluate the effects of NCG and ARG on GT1-7 cell gonadotrophin-releasing hormone (GnRH) secretion, gene expression and cell proliferation. GT1-7 cells were treated in vitro with different concentrations of NCG (0-1.0mM) or ARG (0-4.0mM) in serum-free medium for 12 or 24h. For GnRH secretion and cell proliferation, GT1-7 cells were more sensitive to NCG than ARG...
November 10, 2017: Reproduction, Fertility, and Development
https://www.readbyqxmd.com/read/29109798/multiplexed-nucleic-acid-programmable-protein-arrays
#5
Xiaobo Yu, Lusheng Song, Brianne Petritis, Xiaofang Bian, Haoyu Wang, Jennifer Viloria, Jin Park, Hoang Bui, Han Li, Jie Wang, Lei Liu, Liuhui Yang, Hu Duan, David N McMurray, Jacqueline M Achkar, Mitch Magee, Ji Qiu, Joshua LaBaer
Rationale: Cell-free protein microarrays display naturally-folded proteins based on just-in-time in situ synthesis, and have made important contributions to basic and translational research. However, the risk of spot-to-spot cross-talk from protein diffusion during expression has limited the feature density of these arrays. Methods: In this work, we developed the Multiplexed Nucleic Acid Programmable Protein Array (M-NAPPA), which significantly increases the number of displayed proteins by multiplexing as many as five different gene plasmids within a printed spot...
2017: Theranostics
https://www.readbyqxmd.com/read/29103348/generation-of-thermostable-moloney-murine-leukemia-virus-reverse-transcriptase-variants-using-site-saturation-mutagenesis-library-and-cell-free-protein-expression-system
#6
Yuta Katano, Tongyang Li, Misato Baba, Miyo Nakamura, Masaaki Ito, Kenji Kojima, Teisuke Takita, Kiyoshi Yasukawa
We attempted to increase the thermostability of Moloney murine leukemia virus (MMLV) reverse transcriptase (RT). The eight-site saturation mutagenesis libraries corresponding to Ala70-Arg469 in the whole MMLV RT (Thr24-Leu671), in each of which 1 out of 50 amino acid residues was replaced with other amino acid residue, were constructed. Seven-hundred and sixty eight MMLV RT clones were expressed using a cell-free protein expression system, and their thermostabilities were assessed by the temperature of thermal treatment at which they retained cDNA synthesis activity...
November 6, 2017: Bioscience, Biotechnology, and Biochemistry
https://www.readbyqxmd.com/read/29097185/serum-starvation-of-arpe-19-changes-the-cellular-distribution-of-cholesterol-and-fibulin3-in-patterns-reminiscent-of-age-related-macular-degeneration
#7
Dinusha Rajapakse, Katherine Peterson, Sanghamitra Mishra, Graeme Wistow
Retinal pigment epithelium (RPE) has been implicated as key source of cholesterol-rich deposits at Bruch's membrane (BrM) and in drusen in aging human eye. We have shown that serum-deprivation of confluent RPE cells is associated with upregulation of cholesterol synthesis and accumulation of unesterified cholesterol (UC). Here we investigate the cellular processes involved in this response. We compared the distribution and localization of UC and esterified cholesterol (EC); the age-related macular degeneration (AMD) associated EFEMP1/Fibulin3 (Fib3); and levels of acyl-coenzyme A (CoA): cholesterol acyltransferases (ACAT) ACAT1, ACAT2 and Apolipoprotein B (ApoB) in ARPE-19 cells cultured in serum-supplemented and serum-free media...
October 30, 2017: Experimental Cell Research
https://www.readbyqxmd.com/read/29095525/trichostatin-a-alters-cytoskeleton-and-energy-metabolism-of-pancreatic-adenocarcinoma-cells-an-in-depth-proteomic-study
#8
Elisa Dalla Pozza, Marcello Manfredi, Jessica Brandi, Arianna Buzzi, Eleonora Conte, Raffaella Pacchiana, Daniela Cecconi, Emilio Marengo, Massimo Donadelli
Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal of all human cancers with a high mortality rate. Resistance to conventional treatments and chemotherapeutics is a typical feature of PDAC. To investigate the causes of drug resistance it is essential to deeply investigate the mechanism of action of chemotherapeutics. In this study, we performed an in depth shotgun proteomic approach using the label-free proteomic SWATH-MS analysis to investigate novel insights of the mechanism of action of the histone deacetylase (HDAC) inhibitor trichostatin A (TSA) in PDAC cells...
November 2, 2017: Journal of Cellular Biochemistry
https://www.readbyqxmd.com/read/29090639/platelet-rich-concentrate-in-serum-free-medium-enhances-cartilage-specific-extracellular-matrix-synthesis-and-reduces-chondrocyte-hypertrophy-of-human-mesenchymal-stromal-cells-encapsulated-in-alginate
#9
Shani Samuel, Raja Elina Ahmad, Thamil Selvee Ramasamy, Puvanan Karunanithi, Sangeetha Vasudevaraj Naveen, Tunku Kamarul
Platelet-rich concentrate (PRC), used in conjunction with other chondroinductive growth factors, have been shown to induce chondrogenesis of human mesenchymal stromal cells (hMSC) in pellet culture. However, pellet culture systems promote cell hypertrophy and the presence of other chondroinductive growth factors in the culture media used in previous studies obscures accurate determination of the effect of platelet itself in inducing chondrogenic differentiation. Hence, this study aimed to investigate the effect of PRC alone in enhancing the chondrogenic differentiation potential of human mesenchymal stromal cells (hMSC) encapsulated in three-dimensional alginate constructs...
November 1, 2017: Platelets
https://www.readbyqxmd.com/read/29075041/deregulated-expression-and-activity-of-farnesyl-diphosphate-synthase-fdps-in-glioblastoma
#10
Mario Abate, Chiara Laezza, Simona Pisanti, Giovanni Torelli, Vincenzo Seneca, Giuseppe Catapano, Francesco Montella, Roberta Ranieri, Maria Notarnicola, Patrizia Gazzerro, Maurizio Bifulco, Elena Ciaglia
Glioblastoma (GBM), the most aggressive brain cancer, is highly dependent on the mevalonate (MVA) pathway for the synthesis of lipid moieties critical for cell proliferation but the function and regulation of key intermediate enzymes like farnesyl-diphosphate synthase (FDPS), up to now, remained unknown. A deregulated expression and activity of FDPS was the central research idea of the present study. FDPS mRNA, protein and enzyme activity were analyzed in a cohort of stage III-IV glioma patients (N = 49) and primary derived cells...
October 26, 2017: Scientific Reports
https://www.readbyqxmd.com/read/29070795/ecobody-technology-rapid-monoclonal-antibody-screening-method-from-single-b-cells-using-cell-free-protein-synthesis-for-antigen-binding-fragment-formation
#11
Teruyo Ojima-Kato, Satomi Nagai, Hideo Nakano
We report a rapid and cost-effective monoclonal antibody screening method from single animal B cells using reverse transcription (RT)-PCR and Escherichia coli cell-free protein synthesis (CFPS), which allows evaluation of antibodies within 2 working days. This process is named "Ecobody technology". The method includes strategies to isolate B cells that specifically bind an antigen from the peripheral blood of immunised animals, and single-cell RT-PCR to generate DNA fragments of the VH and VL genes, followed by CFPS for production of fragments of antigen binding (Fab)...
October 25, 2017: Scientific Reports
https://www.readbyqxmd.com/read/29062966/bacterial-cell-free-expression-technology-to-in%C3%A2-vitro-systems-engineering-and-optimization
#12
REVIEW
Filippo Caschera
Cell-free expression system is a technology for the synthesis of proteins in vitro. The system is a platform for several bioengineering projects, e.g. cell-free metabolic engineering, evolutionary design of experiments, and synthetic minimal cell construction. Bacterial cell-free protein synthesis system (CFPS) is a robust tool for synthetic biology. The bacteria lysate, the DNA, and the energy module, which are the three optimized sub-systems for in vitro protein synthesis, compose the integrated system...
June 2017: Synthetic and Systems Biotechnology
https://www.readbyqxmd.com/read/29062958/cell-free-synthetic-biology-engineering-in-an-open-world
#13
REVIEW
Yuan Lu
Cell-free synthetic biology emerges as a powerful and flexible enabling technology that can engineer biological parts and systems for life science applications without using living cells. It provides simpler and faster engineering solutions with an unprecedented freedom of design in an open environment than cell system. This review focuses on recent developments of cell-free synthetic biology on biological engineering fields at molecular and cellular levels, including protein engineering, metabolic engineering, and artificial cell engineering...
March 2017: Synthetic and Systems Biotechnology
https://www.readbyqxmd.com/read/29058591/proteomic-analysis-of-trochophore-and-veliger-larvae-development-in-the-small-abalone-haliotis-diversicolor
#14
Guilan Di, Xianghui Kong, Xiulian Miao, Yifang Zhang, Miaoqin Huang, Yuting Gu, Weiwei You, Jianxin Zhang, Caihuan Ke
BACKGROUND: Haliotis diversicolor is commercially important species. The trochophore and veliger are distinct larval stages in gastropod development. Their development involves complex morphological and physiological changes. We studied protein changes during the embryonic development of H. diversicolor using two dimensional electrophoresis (2-DE) and label-free methods, tandem mass spectrometry (MS/ MS), and Mascot for protein identification. RESULTS: A total of 150 2-DE gel spots were identified...
October 23, 2017: BMC Genomics
https://www.readbyqxmd.com/read/29052140/prion-protein-family-contributes-to-tumorigenesis-via-multiple-pathways
#15
Xiaowen Yang, Zhijun Cheng, Lihua Zhang, Guiru Wu, Run Shi, Zhenxing Gao, Chaoyang Li
A wealth of evidence suggests that proteins from prion protein (PrP) family contribute to tumorigenesis in many types of cancers, including pancreatic ductal adenocarcinoma (PDAC), breast cancer, glioblastoma, colorectal cancer, gastric cancer, melanoma, etc. It is well documented that PrP is a biomarker for PDAC, breast cancer, and gastric cancer. However, the underlying mechanisms remain unclear. The major reasons for cancer cell-caused patient death are metastasis and multiple drug resistance, both of which connect to physiological functions of PrP expressing in cancer cells...
2017: Advances in Experimental Medicine and Biology
https://www.readbyqxmd.com/read/29046485/dimethyl-fumarate-increases-fetal-hemoglobin-provides-heme-detoxification-and-corrects-anemia-in-sickle-cell-disease
#16
Sriram Krishnamoorthy, Betty Pace, Dipti Gupta, Sarah Sturtevant, Biaoru Li, Levi Makala, Julia Brittain, Nancy Moore, Benjamin F Vieira, Timothy Thullen, Ivan Stone, Huo Li, William E Hobbs, David R Light
Sickle cell disease (SCD) results from a point mutation in the β-globin gene forming hemoglobin S (HbS), which polymerizes in deoxygenated erythrocytes, triggering recurrent painful vaso-occlusive crises and chronic hemolytic anemia. Reactivation of fetal Hb (HbF) expression ameliorates these symptoms of SCD. Nuclear factor (erythroid derived-2)-like 2 (Nrf2) is a transcription factor that triggers cytoprotective and antioxidant pathways to limit oxidative damage and inflammation and increases HbF synthesis in CD34+ stem cell-derived erythroid progenitors...
October 19, 2017: JCI Insight
https://www.readbyqxmd.com/read/29043713/bioorthogonal-chemical-imaging-of-metabolic-changes-during-epithelial-mesenchymal-transition-of-cancer-cells-by-stimulated-raman-scattering-microscopy
#17
Luyuan Zhang, Wei Min
Study of metabolic changes during epithelial-mesenchymal transition (EMT) of cancer cells is important for basic understanding and therapeutic management of cancer progression. We here used metabolic labeling and stimulated Raman scattering (SRS) microscopy, a strategy of bioorthogonal chemical imaging, to directly visualize changes in anabolic metabolism during cancer EMT at a single-cell level. MCF-7 breast cancer cell is employed as a model system. Four types of metabolites (amino acids, glucose, fatty acids, and choline) are labeled with either deuterium or alkyne (C≡C) tag...
October 2017: Journal of Biomedical Optics
https://www.readbyqxmd.com/read/29040838/nutrient-sensing-and-metabolic-changes-after-methionine-deprivation-in-primary-muscle-cells-of-turbot-scophthalmus-maximus-l
#18
Haowen Jiang, Fuyun Bian, Huihui Zhou, Xuan Wang, Kaidi Wang, Kangsen Mai, Gen He
The low methionine content in plant-based diets is a major limiting factor for feed utilization by animals. However, the molecular consequences triggered by methionine deficiency have not been well characterized, especially in fish species, whose metabolism is unique in many aspects and important for aquaculture industry. In the present study, the primary muscle cells of turbot (Scophthalmus maximus L.) were isolated and treated with or without methionine for 12 h in culture. The responses of nutrient sensing pathways, the proteomic profiling of metabolic processes, and the expressions of key metabolic molecules were systematically examined...
September 14, 2017: Journal of Nutritional Biochemistry
https://www.readbyqxmd.com/read/29039646/proteomics-based-tools-for-evaluation-of-cell-free-protein-synthesis
#19
Gregory B Hurst, Keiji G Asano, Charles J Doktycz, Elliot J Consoli, William L Doktycz, Carmen M Foster, Jennifer L Morrell-Falvey, Robert F Standaert, Mitchel J Doktycz
Cell-free protein synthesis (CFPS) has the potential to produce enzymes, therapeutic agents, and other proteins, while circumventing difficulties associated with in vivo heterologous expression. However, the contents of the cell-free extracts used to carry out synthesis are generally not characterized, which hampers progress toward enhancing yield or functional activity of the target protein. We explored the utility of mass spectrometry (MS)-based proteomics for characterizing the bacterial extracts used for transcribing and translating gene sequences into proteins as well as the products of CFPS reactions...
November 7, 2017: Analytical Chemistry
https://www.readbyqxmd.com/read/29034959/quantitative-proteomics-of-a-b12-dependent-alga-grown-in-coculture-with-bacteria-reveals-metabolic-tradeoffs-required-for-mutualism
#20
Katherine E Helliwell, Jagroop Pandhal, Matthew B Cooper, Joseph Longworth, Ulrich Johan Kudahl, David A Russo, Eleanor V Tomsett, Freddy Bunbury, Deborah L Salmon, Nicholas Smirnoff, Phillip C Wright, Alison G Smith
The unicellular green alga Lobomonas rostrata requires an external supply of vitamin B12 (cobalamin) for growth, which it can obtain in stable laboratory cultures from the soil bacterium Mesorhizobium loti in exchange for photosynthate. We investigated changes in protein expression in the alga that allow it to engage in this mutualism. We used quantitative isobaric tagging (iTRAQ) proteomics to determine the L. rostrata proteome grown axenically with B12 supplementation or in coculture with M. loti. Data are available via ProteomeXchange (PXD005046)...
October 16, 2017: New Phytologist
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