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membrane protein purification

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https://www.readbyqxmd.com/read/28526409/expression-and-purification-of-functional-pdgf-receptor-beta
#1
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/28518108/identification-of-plant-ice-binding-proteins-through-assessment-of-ice-recrystallization-inhibition-and-isolation-using-ice-affinity-purification
#2
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/28470611/expression-of-prokaryotic-integral-membrane-proteins-in-e-coli
#3
James D Love
Production of prokaryotic membrane proteins for structural and functional studies in E. coli can be parallelized and miniaturized. All stages from cloning, expression, purification to detergent selection can be investigated using high-throughput techniques to rapidly and economically find tractable targets.
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28469466/an-improved-2-dimensional-gel-electrophoresis-method-for-resolving-human-erythrocyte-membrane-proteins
#4
Manoj Kumar, Rajendra Singh, Anil Meena, Bhagwan S Patidar, Rajendra Prasad, Sunil K Chhabra, Surendra K Bansal
The 2-dimensional gel electrophoresis (2-DE) technique is widely used for the analysis of complex protein mixtures extracted from biological samples. It is one of the most commonly used analytical techniques in proteomics to study qualitative and quantitative protein changes between different states of a cell or an organism (eg, healthy and diseased), conditionally expressed proteins, posttranslational modifications, and so on. The 2-DE technique is used for its unparalleled ability to separate thousands of proteins simultaneously...
2017: Proteomic Insights
https://www.readbyqxmd.com/read/28465586/oxidation-of-protein-bound-methionine-in-photofrin-photodynamic-therapy-treated-human-tumor-cells-explored-by-methionine-containing-peptide-enrichment-and-quantitative-proteomics-approach
#5
Ya-Ju Hsieh, Kun-Yi Chien, I-Fang Yang, I-Neng Lee, Chia-Chun Wu, Tung-Yung Huang, Jau-Song Yu
In Photofrin-mediated photodynamic therapy (PDT), cell fate can be modulated by the subcellular location of Photofrin. PDT triggers oxidative damage to target cells, including the methionine (Met) oxidation of proteins. Here, we developed a new Met-containing peptide enrichment protocol combined with SILAC-based quantitative proteomics, and used this approach to explore the global Met oxidation changes of proteins in PDT-treated epidermoid carcinoma A431 cells preloaded with Photofrin at the plasma membrane, ER/Golgi, or ubiquitously...
May 2, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28454841/assisted-and-unassisted-protein-insertion-into-liposomes
#6
REVIEW
Andreas Kuhn, Maximilian Haase, Sebastian Leptihn
The insertion of newly synthesized membrane proteins is a well-regulated and fascinating process occurring in every living cell. Several translocases and insertases have been found in prokaryotic and eukaryotic cells, the Sec61 complex and the Get complex in the endoplasmic reticulum and the SecYEG complex and YidC in bacteria and archaea. In mitochondria, TOM and TIM complexes transport nuclear-encoded proteins, whereas the Oxa1 is required for the insertion of mitochondria-encoded membrane proteins. Related to the bacterial YidC and the mitochondrial Oxa1 are the Alb3 and Alb4 proteins in chloroplasts...
April 25, 2017: Biophysical Journal
https://www.readbyqxmd.com/read/28452363/mechanism-of-sos-pr-domain-autoinhibition-revealed-by-single-molecule-assays-on-native-protein-from-lysate
#7
Young Kwang Lee, Shalini T Low-Nam, Jean K Chung, Scott D Hansen, Hiu Yue Monatrice Lam, Steven Alvarez, Jay T Groves
The guanine nucleotide exchange factor (GEF) Son of Sevenless (SOS) plays a critical role in signal transduction by activating Ras. Here we introduce a single-molecule assay in which individual SOS molecules are captured from raw cell lysate using Ras-functionalized supported membrane microarrays. This enables characterization of the full-length SOS protein, which has not previously been studied in reconstitution due to difficulties in purification. Our measurements on the full-length protein reveal a distinct role of the C-terminal proline-rich (PR) domain to obstruct the engagement of allosteric Ras independently of the well-known N-terminal domain autoinhibition...
April 28, 2017: Nature Communications
https://www.readbyqxmd.com/read/28435877/design-and-function-of-biomimetic-multilayer-water-purification-membranes
#8
Shengjie Ling, Zhao Qin, Wenwen Huang, Sufeng Cao, David L Kaplan, Markus J Buehler
Multilayer architectures in water purification membranes enable increased water throughput, high filter efficiency, and high molecular loading capacity. However, the preparation of membranes with well-organized multilayer structures, starting from the nanoscale to maximize filtration efficiency, remains a challenge. We report a complete strategy to fully realize a novel biomaterial-based multilayer nanoporous membrane via the integration of computational simulation and experimental fabrication. Our comparative computational simulations, based on coarse-grained models of protein nanofibrils and mineral plates, reveal that the multilayer structure can only form with weak interactions between nanofibrils and mineral plates...
April 2017: Science Advances
https://www.readbyqxmd.com/read/28431999/novel-method-to-rapidly-and-efficiently-lyse-escherichia-coli-for-the-isolation-of-recombinant-protein
#9
Himanshu Joshi, Vikas Jain
Rapid and high-throughput protein purification methods are required to explore structure and function of several uncharacterized proteins. Isolation of recombinant protein expressed in Escherichia coli strain BL21 (DE3) depends largely on the efficient and speedy bacterial cell lysis, which is considered as the bottleneck during protein purification. Cells are usually lysed by either sonication or high pressure homogenization, both of which are slow, require special equipment, lead to heat generation, and may result in loss of protein's biological activity...
April 18, 2017: Analytical Biochemistry
https://www.readbyqxmd.com/read/28417934/expression-of-adenosine-a2b-receptor-and-adenosine-deaminase-in-rabbit-gastric-mucosa-ecl-cells
#10
Rosa María Arin, Ana Isabel Vallejo, Yuri Rueda, Olatz Fresnedo, Begoña Ochoa
Adenosine is readily available to the glandular epithelium of the stomach. Formed continuously in intracellular and extracellular locations, it is notably produced from ATP released in enteric cotransmission. Adenosine analogs modulate chloride secretion in gastric glands and activate acid secretion in isolated parietal cells through A2B adenosine receptor (A2BR) binding. A functional link between surface A2BR and adenosine deaminase (ADA) was found in parietal cells, but whether this connection is a general feature of gastric mucosa cells is unknown...
April 12, 2017: Molecules: a Journal of Synthetic Chemistry and Natural Product Chemistry
https://www.readbyqxmd.com/read/28413443/molecular-and-biochemical-characterization-of-recombinant-cel12b-cel8c-and-peh28-overexpressed-in-escherichia-coli-and-their-potential-in-biofuel-production
#11
Eman Ibrahim, Kim D Jones, Keith E Taylor, Ebtesam N Hosseney, Patrick L Mills, Jean M Escudero
BACKGROUND: The high crystallinity of cellulosic biomass myofibrils as well as the complexity of their intermolecular structure is a significant impediment for biofuel production. Cloning of celB-, celC-encoded cellulases (cel12B and cel8C) and peh-encoded polygalacturonase (peh28) from Pectobacterium carotovorum subsp. carotovorum (Pcc) was carried out in our previous study using Escherichia coli as a host vector. The current study partially characterizes the enzymes' molecular structures as well as their catalytic performance on different substrates which can be used to improve their potential for lignocellulosic biomass conversion...
2017: Biotechnology for Biofuels
https://www.readbyqxmd.com/read/28409449/isolation-of-native-soluble-and-membrane-bound-protein-complexes-from-yeast-saccharomyces-cerevisiae
#12
Tobias Hansen, Anna Chan, Thomas Schröter, Daniel Schwerter, Wolfgang Girzalsky, Ralf Erdmann
Immunoprecipitation is a traditional approach to isolate single proteins or native protein complexes from a complex sample mixture. The original method makes use of specific antibodies against endogenous proteins or epitope tags, which are first bound to the target protein and then isolated with protein A beads. An advancement of this method is the application of a protein A tag fused to the target protein and the affinity-purification of the tagged protein with human Immunoglobulin G chemically cross-linked to a sepharose matrix...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28402172/rational-design-and-synthesis-of-affinity-matrices-based-on-proteases-immobilized-onto-cellulose-membranes
#13
Alberto Del Monte-Martínez, Jorge González-Bacerio, Bessy Cutiño-Avila, Jorge Rojas, Mae Chappé, Emir Salas-Sarduy, Isel Pascual, José M Guisán
Discovery of new protease inhibitors may result in potential therapeutic agents or useful biotechnological tools. Obtainment of these molecules from natural sources requires simple, economic and highly efficient purification protocols. The aim of this work was the obtainment of affinity matrices by the covalent immobilization of dipeptidyl peptidase IV (DPP-IV) and papain onto cellulose membranes, previously activated with formyl (FCM) or glyoxyl groups (GCM). GCM showed the highest activation grade (10.2 µmol aldehyde/cm(2))...
April 12, 2017: Preparative Biochemistry & Biotechnology
https://www.readbyqxmd.com/read/28399395/spectraplakin-induces-positive-feedback-between-fusogens-and-the-actin-cytoskeleton-to-promote-cell-cell-fusion
#14
Yihong Yang, Yan Zhang, Wen-Jun Li, Yuxiang Jiang, Zhiwen Zhu, Huifang Hu, Wei Li, Jia-Wei Wu, Zhi-Xin Wang, Meng-Qiu Dong, Shanjin Huang, Guangshuo Ou
Cell-cell fusion generally requires cellular fusogenic proteins and actin-propelled membrane protrusions. However, the molecular connections between fusogens and the actin cytoskeleton remain unclear. Here, we show that the Caenorhabditis elegans fusogen EFF-1 and F-actin are enriched at the cortex of the post-embryonic fusing cells, and conditional mutations of WASP and Arp2/3 delay cell-cell fusion by impairing EFF-1 localization. Our affinity purification and mass spectrometry analyses determined that an actin-binding protein, spectraplakin/VAB-10A, binds to EFF-1...
April 10, 2017: Developmental Cell
https://www.readbyqxmd.com/read/28380693/a-non-chromatographic-process-for-purification-of-secretory-immunoglobulins-from-caprine-whey
#15
Alexander Matlschweiger, Gottfried Himmler, Clemens Linhart, Michael Harasek, Rainer Hahn
Secretory immunoglobulins are an important antibody class being primarily responsible for immunoprotection of mucosal surfaces. A simple, non-chromatographic purification process for secretory immunoglobulins from caprine whey was developed. In the first process step whey was concentrated 30 - 40 fold on a 500 kDa membrane, thereby increasing the purity from 3% to 15%. The second step consisted of a fractionated PEG precipitation, in which high molecular weight impurities were removed first and in the second stage the secretory immunoglobulins were precipitated, leaving a majority of the low molecular weight proteins in solution...
April 5, 2017: Biotechnology Progress
https://www.readbyqxmd.com/read/28374993/removable-backbone-modification-method-for-the-chemical-synthesis-of-membrane-proteins
#16
Jia-Bin Li, Shan Tang, Ji-Shen Zheng, Chang-Lin Tian, Lei Liu
Chemical synthesis can produce water-soluble globular proteins bearing specifically designed modifications. These synthetic molecules have been used to study the biological functions of proteins and to improve the pharmacological properties of protein drugs. However, the above advances notwithstanding, membrane proteins (MPs), which comprise 20-30% of all proteins in the proteomes of most eukaryotic cells, remain elusive with regard to chemical synthesis. This difficulty stems from the strong hydrophobic character of MPs, which can cause considerable handling issues during ligation, purification, and characterization steps...
April 4, 2017: Accounts of Chemical Research
https://www.readbyqxmd.com/read/28366668/the-intermembrane-space-protein-erv1-of-trypanosoma-brucei-is-essential-for-mitochondrial-fe-s-cluster-assembly-and-operates-alone
#17
Alexander C Haindrich, Michala Boudová, Marie Vancová, Priscila Peña Diaz, Eva Horáková, Julius Lukeš
Sulfhydryl oxidase Erv1 is a ubiquitous and conserved protein of the mitochondrial intermembrane space that plays a role in the transport of small sulfur-containing proteins. In higher eukaryotes, Erv1 interacts with the mitochondrial import protein Mia40. However, Trypanosoma brucei lacks an obvious Mia40 homologue in its genome. Here we show by tandem affinity purification and mass spectrometry that in this excavate protist, Erv1 functions without a Mia40 homologue and most likely any other interaction partner...
March 31, 2017: Molecular and Biochemical Parasitology
https://www.readbyqxmd.com/read/28364041/galectin-3-interacts-with-the-cell-surface-glycoprotein-cd146-mcam-muc18-and-induces-secretion-of-metastasis-promoting-cytokines-from-vascular-endothelial-cells
#18
Florent Colomb, Weikun Wang, Deborah Simpson, Mudaser Zafar, Robert Beynon, Jonathan M Rhodes, Lu-Gang Yu
The galactoside-binding protein galectin-3 is increasingly recognized as an important player in cancer development, progression, and metastasis via its interactions with various galactoside-terminated glycans. We have previously shown that circulating galectin-3, that is increased by up to 30-fold in cancer patients, promotes blood-borne metastasis in an animal cancer model. This effect is partly attributable to galectin-3 interaction with unknown receptor(s) on vascular endothelial cells and causes endothelial secretion of several metastasis-promoting cytokines...
March 31, 2017: Journal of Biological Chemistry
https://www.readbyqxmd.com/read/28343687/multiple-simultaneous-independent-gradients-for-a-versatile-multidimensional-liquid-chromatography-part-ii-application-2-computer-controlled-ph-gradients-in-the-presence-of-urea-provide-improved-separation-of-proteins-stability-influenced-anion-and-cation-exchange
#19
Allen G Hirsh, Latchezar I Tsonev
This paper details the use of a method of creating controlled pH gradients (pISep) to improve the separation of protein isoforms on ion exchange (IEX) stationary phases in the presence of various isocratic levels of urea. The pISep technology enables the development of computer controlled pH gradients on both cationic (CEX) and anionic (AEX) IEX stationary phases over the very wide pH range from 2 to 12. In pISep, titration curves generated by proportional mixing of the acidic and basic pISep working buffers alone, or in the presence of non-buffering solutes such as the neutral salt NaCl (0-1M), polar organics such as urea (0-8M) or acetonitrile (0-80 Vol%), can be fitted with high fidelity using high order polynomials which, in turn allows construction of a mathematical manifold %A (% acidic pISep buffer) vs...
April 28, 2017: Journal of Chromatography. A
https://www.readbyqxmd.com/read/28340620/stable-high-level-expression-of-factor-viii-in-chinese-hamster-ovary-cells-in-improved-elongation-factor-1-alpha-based-system
#20
Nadezhda A Orlova, Sergey V Kovnir, Alexandre G Gabibov, Ivan I Vorobiev
BACKGROUND: Recombinant factor VIII (FVIII), used for haemophilia A therapy, is one of the most challenging among the therapeutic proteins produced in heterologous expression systems. Deletion variant of FVIII, in which the entire domain B is replaced by a short linker peptide, was approved for medical use. Efficacy and safety of this FVIII deletion variant are similar to full-length FVIII preparations while the level of production in CHO cells is substantially higher. Typical levels of productivity for CHO cell lines producing deletion variant FVIII-BDD SQ, described elsewhere, are 0...
March 24, 2017: BMC Biotechnology
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