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https://www.readbyqxmd.com/read/28535613/expression-and-purification-of-biologically-active-human-bone-morphogenetic-protein-4-in-recombinant-chinese-hamster-ovary-cells
#1
Minyub Cha, Nara Han, Jia Pi, Yongsu Jeong, Kwanghee Baek, Jaeseung Yoon
Bone morphogenetic protein-4 (BMP-4) is considered to have therapeutic potential for various diseases including cancers; however, the high expression of biologically active recombinant human BMP-4 (rhBMP-4) needed to manufacture rhBMP-4 for therapeutic purposes has yet to be established. In the current study, we established a recombinant Chinese hamster ovary (rCHO) cell line overexpressing rhBMP-4 as well as a production process using 7.5-l bioreactor (5 l working volume). The expression of the mature rhBMP-4 was significantly enhanced by recombinant furin expression...
May 24, 2017: Journal of Microbiology and Biotechnology
https://www.readbyqxmd.com/read/28534782/reinforce-an-ensemble-approach-for-inferring-ppi-network-from-ap-ms-data
#2
Bo Tian, Qiong Duan, Can Zhao, Ben Teng, Zengyou He
Affinity Purification-Mass Spectrometry (AP-MS) is one of the most important technologies for constructing protein-protein interaction (PPI) networks. In this paper, we propose an ensemble method, Reinforce, for inferring PPI network from AP-MS data set. The new algorithm named Reinforce is based on rank aggregation and false discovery rate control. Under the null hypothesis that the interaction scores from different scoring methods are randomly generated, Reinforce follows three steps to integrate multiple ranking results from different algorithms or different data sets...
May 17, 2017: IEEE/ACM Transactions on Computational Biology and Bioinformatics
https://www.readbyqxmd.com/read/28534518/cdk20-interacts-with-keap1-to-activate-nrf2-and-promotes-radiochemoresistance-in-lung-cancer-cells
#3
Q Wang, J Ma, Y Lu, S Zhang, J Huang, J Chen, J-X Bei, K Yang, G Wu, K Huang, J Chen, S Xu
Radiochemoresistance is considered the main cause of local recurrence and distant metastasis in lung cancer. However, the underlying mechanisms of radiochemoresistance remain to be uncovered. In this study, we determine the functions of cell cycle-related kinase (CDK20) in radiochemoresistance. CDK20 is a newly identified protein kinase, which plays critical roles in cell growth and proliferation in several types of cancer. Using tandem affinity purification technology, we provide evidences that CDK20 binds to the ubiquitin ligase Kelch-like ECH-associated protein 1 (KEAP1), which targets transcriptional factor nuclear factor erythroid-2-related factor 2 (NRF2) for degradation...
May 22, 2017: Oncogene
https://www.readbyqxmd.com/read/28534479/efficient-protein-production-inspired-by-how-spiders-make-silk
#4
Nina Kronqvist, Médoune Sarr, Anton Lindqvist, Kerstin Nordling, Martins Otikovs, Luca Venturi, Barbara Pioselli, Pasi Purhonen, Michael Landreh, Henrik Biverstål, Zigmantas Toleikis, Lisa Sjöberg, Carol V Robinson, Nicola Pelizzi, Hans Jörnvall, Hans Hebert, Kristaps Jaudzems, Tore Curstedt, Anna Rising, Jan Johansson
Membrane proteins are targets of most available pharmaceuticals, but they are difficult to produce recombinantly, like many other aggregation-prone proteins. Spiders can produce silk proteins at huge concentrations by sequestering their aggregation-prone regions in micellar structures, where the very soluble N-terminal domain (NT) forms the shell. We hypothesize that fusion to NT could similarly solubilize non-spidroin proteins, and design a charge-reversed mutant (NT*) that is pH insensitive, stabilized and hypersoluble compared to wild-type NT...
May 23, 2017: Nature Communications
https://www.readbyqxmd.com/read/28529978/emerging-applications-of-exosomes-in-cancer-therapeutics-and-diagnostics
#5
REVIEW
Sahil Inamdar, Rajeshwar Nitiyanandan, Kaushal Rege
Exosomes are nanoscale extracellular vesicles that are shed from different cells in the body. Exosomes encapsulate several biomolecules including lipids, proteins, and nucleic acids, and can therefore play a key role in cellular communication. These vesicles can be isolated from different body fluids and their small sizes make them attractive in various biomedical applications. Here, we review state-of-the art approaches in exosome isolation and purification, and describe their potential use in cancer vaccines, drug delivery, and diagnostics...
March 2017: Bioengineering & Translational Medicine
https://www.readbyqxmd.com/read/28529565/prodelphinidins-isolated-from-chinese-bayberry-leaves-induces-apoptosis-via-the-p53-dependent-signaling-pathways-in-ovcar-3-human-ovarian-cancer-cells
#6
Yu Fu, Xingqian Ye, Malcolm Lee, Gary Rankin, Yi Charlie Chen
Chinese bayberry leaves are rich in prodelphinidins. Since the isolation and purification of prodelphinidins is difficult, the association between the degree of prodelphinidin polymerization and their anti-carcinogenic activity remains ambiguous. The cytotoxic and apoptotic activities of prodelphinidin Chinese bayberry leaf extracts (PCBLs), oligomeric proanthocyanidins (OPAs) and polymeric proanthocyanidins (PPAs), isolated by normal-phase preparative high-performance liquid chromatography were investigated in OVCAR-3 human ovarian cancer cells...
May 2017: Oncology Letters
https://www.readbyqxmd.com/read/28526525/high-throughput-estimation-of-specific-activities-of-enzyme-mutants-in-cell-lysates-through-immunoturbidimetric-assay-of-proteins
#7
Xiaolan Yang, Yiran Feng, Huimin Chong, Deqiang Wang, Xiaolei Hu, Jun Pu, Chang-Guo Zhan, Fei Liao
High-throughput estimation of specific activities of an enzyme and its mutants in a group (enzyme/mutants) in cell lysates via high-throughput assay of their activities and separate immunoturbidimetric assay (ITA) of their proteins was proposed. Pseudomonas aeruginosa arylsulfatase (PAAS) and Bacillus fastidious uricase (BFU) served as two models. ITA employed 0.75 mg of antisera against PAAS or BFU as the reference in 96-well microplates to measure the difference of extinction at 340 and 700 nm. According to the calibration curve, ITA quantified the reference from 0...
May 16, 2017: Analytical Biochemistry
https://www.readbyqxmd.com/read/28526409/expression-and-purification-of-functional-pdgf-receptor-beta
#8
Qingbin Shang, Liang Zhao, Xiaojing Wang, Meimei Wang, Sen-Fang Sui, Li-Zhi Mi
Platelet Derived Growth Factor receptors (PDGFRs), members of receptor tyrosine kinase superfamily, play essential roles in early hematopoiesis, angiogenesis and organ development. Dysregulation of PDGF receptor signaling under pathological conditions associates with cancers, vascular diseases, and fibrotic diseases. Therefore, they are attractive targets in drug development. Like any other membrane proteins with a single-pass transmembrane domain, the high-resolution structural information of the full-length PDGF receptors is still not resolved...
May 16, 2017: Biochemical and Biophysical Research Communications
https://www.readbyqxmd.com/read/28523998/uncovering-drug-mechanism-of-action-by-proteome-wide-identification-of-drug-binding-proteins
#9
Tao Cui, Hai Hou, Yao Sun, Huaixing Cang, Xiangxi Wang
Small molecule drugs obtained from synthetic compounds or natural products show their therapeutic effects by specifically binding to one or a few target proteins and modulating their functions. In contrast, undesirable drug-protein interactions may provoke harmful side effects. Furthermore, drug-protein interactions also play roles in drug activation, transport, metabolism and regulation of drug resistance. Therefore, systematically identifying binding proteins of a drug molecule is critical for understanding the mechanism of action of the drug at molecular level...
May 18, 2017: Medicinal Chemistry
https://www.readbyqxmd.com/read/28523846/a-novel-rapid-analysis-using-mass-spectrometry-to-evaluate-downstream-refolding-of-recombinant-human-insulin-like-growth-factor-1-mecasermin
#10
Kenichiro Furuki, Toshimasa Toyo'oka, Hideto Yamaguchi
RATIONALE: Mecasermin is used to treat elevated blood sugar as well as growth hormone-resistant Laron-type dwarfism. Mecasermin isolated from inclusion bodies in extracts of E.coli must be refolded to acquire sufficient activity. However, there is no rapid analytical method for monitoring refolding during the purification process. METHODS: We prepared mecasermin drug product, in-process samples during the oxidation of mecasermin, forced-reduced mecasermin, and aerially oxidized mecasermin after forced reduction...
May 19, 2017: Rapid Communications in Mass Spectrometry: RCM
https://www.readbyqxmd.com/read/28522394/interaction-research-on-an-antiviral-molecule-that-targets-the-coat-protein-of-southern-rice-black-streaked-dwarf-virus
#11
Longlu Ran, Yan Ding, Liangzhi Luo, Xiuhai Gan, Xiangyang Li, Yongzhong Chen, Deyu Hu, Baoan Song
Southern rice black-streaked dwarf virus (SRBSDV) coat protein (P10) is the key protein required for viral transmission and host plant infection and is thus a promising target for anti-SRBSDV agent screening. In this study, P10 was obtained from Escherichia coli through cloning, expression, and purification. The antiviral agent Ningnanmycin was selected as control, and a series of antiviral compounds based on the structural scaffold of ferulic acid were analyzed. Size-exclusion chromatography analysis results showed that compound F27 can alter the aggregation of P10 proteins...
May 15, 2017: International Journal of Biological Macromolecules
https://www.readbyqxmd.com/read/28518108/identification-of-plant-ice-binding-proteins-through-assessment-of-ice-recrystallization-inhibition-and-isolation-using-ice-affinity-purification
#12
Melissa Bredow, Heather E Tomalty, Virginia K Walker
Ice-binding proteins (IBPs) belong to a family of stress-induced proteins that are synthesized by certain organisms exposed to subzero temperatures. In plants, freeze damage occurs when extracellular ice crystals grow, resulting in the rupture of plasma membranes and possible cell death. Adsorption of IBPs to ice crystals restricts further growth by a process known as ice-recrystallization inhibition (IRI), thereby reducing cellular damage. IBPs also demonstrate the ability to depress the freezing point of a solution below the equilibrium melting point, a property known as thermal hysteresis (TH) activity...
May 5, 2017: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/28518087/multimer-page-a-method-for-capturing-and-resolving-protein-complexes-in-biological-samples
#13
Tyler Rhinesmith, Bryan A Killinger, Akhil Sharma, Anna Moszczynska
There are many well-developed methods for purifying and studying single proteins and peptides. However, most cellular functions are carried out by networks of interacting protein complexes, which are often difficult to investigate because their binding is non-covalent and easily perturbed by purification techniques. This work describes a method of stabilizing and separating native protein complexes from unmodified tissue using two-dimensional polyacrylamide gel electrophoresis. Tissue lysate is loaded onto a non-denaturing blue-native polyacrylamide gel, then an electric current is applied until the protein migrates a short distance into the gel...
May 5, 2017: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/28517015/genetic-incorporation-of-4-fluorohistidine-into-peptides-enables-selective-affinity-purification
#14
Christine M Ring, Emil S Iqbal, David E Hacker, Matthew C T Hartman, T Ashton Cropp
Due to the lowered pKa of 4-fluorohistidine relative to histidine, peptides and proteins containing this amino acid are potentially endowed with novel properties. We report here the optimized synthesis of 4-fluorohistidine and show that it can efficiently replace histidine in in vitro translation reactions. Moreover, peptides containing 6×-fluorohistidine tags are able to be selectively captured and eluted from nickel resin in the presence of his-tagged protein mixtures.
May 18, 2017: Organic & Biomolecular Chemistry
https://www.readbyqxmd.com/read/28514442/architecture-of-the-human-interactome-defines-protein-communities-and-disease-networks
#15
Edward L Huttlin, Raphael J Bruckner, Joao A Paulo, Joe R Cannon, Lily Ting, Kurt Baltier, Greg Colby, Fana Gebreab, Melanie P Gygi, Hannah Parzen, John Szpyt, Stanley Tam, Gabriela Zarraga, Laura Pontano-Vaites, Sharan Swarup, Anne E White, Devin K Schweppe, Ramin Rad, Brian K Erickson, Robert A Obar, K G Guruharsha, Kejie Li, Spyros Artavanis-Tsakonas, Steven P Gygi, J Wade Harper
The physiology of a cell can be viewed as the product of thousands of proteins acting in concert to shape the cellular response. Coordination is achieved in part through networks of protein-protein interactions that assemble functionally related proteins into complexes, organelles, and signal transduction pathways. Understanding the architecture of the human proteome has the potential to inform cellular, structural, and evolutionary mechanisms and is critical to elucidating how genome variation contributes to disease...
May 17, 2017: Nature
https://www.readbyqxmd.com/read/28512649/purification-and-characterization-of-schwann-cells-from-adult-human-skin-and-nerve
#16
Jo Anne Stratton, Ranjan Kumar, Sarthak Sinha, Prajay Shah, Morgan Stykel, Yuval Shapira, Rajiv Midha, Jeff Biernaskie
Despite its modest capacity for regeneration, peripheral nervous system injury often results in significant long-term disability. Supplementing peripheral nervous system injury with autologous Schwann cells (SCs) may serve to rejuvenate the postinjury environment to enhance regeneration and ultimately improve functional outcomes. However, human nerve-derived SC (hN-SC) collection procedures require invasive surgical resection. Here, we describe the characterization of SCs from adult human skin (hSk-SCs) of four male donors ranging between 27 and 46 years old...
May 2017: ENeuro
https://www.readbyqxmd.com/read/28512598/phytochemical-screening-and-study-of-antioxidant-antimicrobial-antidiabetic-anti-inflammatory-and-analgesic-activities-of-extracts-from-stem-wood-of-pterocarpus-marsupium-roxburgh
#17
Dipak Raj Pant, Narayan Dutt Pant, Dil Bahadur Saru, Uday Narayan Yadav, Dharma Prasad Khanal
AIMS: The main aims of the study were to evaluate the phytochemical constituents and to study the antioxidant, antimicrobial, antidiabetic, anti-inflammatory, and analgesic activities of extracts from stem wood of Pterocarpus marsupium. MATERIALS AND METHODS: Ethanol, acetone and isopropyl alcohol (IPA) (1:1) extracts of stem wood of P. marsupium were subjected to phytochemical screening and analysis of biological activities from August 2015 to January 2016. The antioxidant assay was carried out using 2, 2-diphenyl-1-picrylhydrazyl scavenging method, antimicrobial activity testing by cup diffusion method, antidiabetic test evaluation by oral glucose tolerance test in mice, anti-inflammatory effect was evaluated by hind paw edema method in mice and analgesic test evaluation by a chemical writhing method in mice...
April 2017: Journal of Intercultural Ethnopharmacology
https://www.readbyqxmd.com/read/28510356/purification-of-recombinant-a%C3%AE-1-42-and-pglu-a%C3%AE-3-42-using-preparative-sds-page
#18
Claudia Spahn, Michael Wermann, Rico Eichentopf, Gerd Hause, Dagmar Schlenzig, Stephan Schilling
Recombinant expression and purification of amyloid peptides represents a common basis for investigating the molecular mechanisms of amyloid formation and toxicity. However, the isolation of the recombinant peptides is hampered by inefficient separation from contaminants such as the fusion protein required for efficient expression in E. coli. Here, we present a new approach for the isolation of highly purified Aβ(1-42) and pGlu-Aβ(3-42), which is based on a separation using preparative SDS-PAGE. The method relies on the purification of the Aβ fusion protein by affinity chromatography followed by preparative SDS-PAGE under reducing conditions and subsequent removal of detergents by precipitation...
May 16, 2017: Electrophoresis
https://www.readbyqxmd.com/read/28509348/reconstitution-of-the-plant-ubiquitination-cascade-in-bacteria-using-a-synthetic-biology-approach
#19
Yufang Han, Jianhang Sun, Jun Yang, Zhaoyun Tan, Jijing Luo, Dongping Lu
Ubiquitination modulates nearly all aspects of plant life. Here, we reconstituted the Arabidopsis thaliana ubiquitination cascade in Escherichia coli using a synthetic biology approach. In this system, plant proteins are expressed and then immediately participate in ubiquitination reactions within E. coli cells. Additionally, the purification of the individual ubiquitination components prior to setting up the ubiquitination reactions is skipped. To establish the reconstituted system, we co-expressed Arabidopsis ubiquitin (Ub) and ubiquitination substrates with E1, E2 and E3 enzymes in E...
May 16, 2017: Plant Journal: for Cell and Molecular Biology
https://www.readbyqxmd.com/read/28508886/protein-protein-interactions-and-metabolite-channelling-in-the-plant-tricarboxylic-acid-cycle
#20
Youjun Zhang, Katherine F M Beard, Corné Swart, Susan Bergmann, Ina Krahnert, Zoran Nikoloski, Alexander Graf, R George Ratcliffe, Lee J Sweetlove, Alisdair R Fernie, Toshihiro Obata
Protein complexes of sequential metabolic enzymes, often termed metabolons, may permit direct channelling of metabolites between the enzymes, providing increased control over metabolic pathway fluxes. Experimental evidence supporting their existence in vivo remains fragmentary. In the present study, we test binary interactions of the proteins constituting the plant tricarboxylic acid (TCA) cycle. We integrate (semi-)quantitative results from affinity purification-mass spectrometry, split-luciferase and yeast-two-hybrid assays to generate a single reliability score for assessing protein-protein interactions...
May 16, 2017: Nature Communications
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