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protein purification

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https://www.readbyqxmd.com/read/28922633/protein-extraction-into-the-bicontinuous-microemulsion-phase-of-a-water-sds-pentanol-dodecane-winsor-iii-system-effect-on-nanostructure-and-protein-conformation
#1
Douglas G Hayes, Ran Ye, Rachel N Dunlap, Matthew J Cuneo, Sai Venkatesh Pingali, Hugh M O'Neill, Volker S Urban
Bicontinuous microemulsions (BμEs), consisting of water and oil nanodomains separated by surfactant monolayers of near-zero curvature, are potentially valuable systems for purification and delivery of biomolecules, for hosting multiphasic biochemical reactions, and as templating media for preparing nanomaterials. We formed Winsor-III systems by mixing aqueous protein and sodium dodecyl sulfate (SDS) solutions with dodecane and 1-pentanol (cosurfactant) to efficiently extract proteins into the middle (BμE) phase...
September 7, 2017: Colloids and Surfaces. B, Biointerfaces
https://www.readbyqxmd.com/read/28922413/exploitation-of-stable-nanostructures-based-on-the-mouse-polyomavirus-for-development-of-a-recombinant-vaccine-against-porcine-circovirus-2
#2
Martin Fraiberk, Michaela Hájková, Magdaléna Krulová, Martina Kojzarová, Alena Drda Morávková, Ivan Pšikal, Jitka Forstová
The aim of this study was to develop a suitable vaccine antigen against porcine circovirus 2 (PCV2), the causative agent of post-weaning multi-systemic wasting syndrome, which causes significant economic losses in swine breeding. Chimeric antigens containing PCV2b Cap protein sequences based on the mouse polyomavirus (MPyV) nanostructures were developed. First, universal vectors for baculovirus-directed production of chimeric MPyV VLPs or pentamers of the major capsid protein, VP1, were designed for their exploitation as vaccines against other pathogens...
2017: PloS One
https://www.readbyqxmd.com/read/28922409/purification-and-characterisation-of-the-yeast-plasma-membrane-atp-binding-cassette-transporter-pdr11p
#3
Katrine Rude Laub, Magdalena Marek, Lyubomir Dimitrov Stanchev, Sara Abad Herrera, Tamara Kanashova, Adèle Bourmaud, Gunnar Dittmar, Thomas Günther Pomorski
The ATP binding cassette (ABC) transporters Pdr11p and its paralog Aus1p are expressed under anaerobic growth conditions at the plasma membrane of the yeast Saccharomyces cerevisiae and are required for sterol uptake. However, the precise mechanism by which these ABC transporters facilitate sterol movement is unknown. In this study, an overexpression and purification procedure was developed with the aim to characterise the Pdr11p transporter. Engineering of Pdr11p variants fused at the C terminus with green fluorescent protein (Pdr11p-GFP) and containing a FLAG tag at the N terminus facilitated expression analysis and one-step purification, respectively...
2017: PloS One
https://www.readbyqxmd.com/read/28921494/solid-phase-synthesis-of-rna-analogs-containing-phosphorodithioate-linkages
#4
Xianbin Yang
The oligoribonucleotide phosphorodithioate (PS2-RNA) modification uses two sulfur atoms to replace two non-bridging oxygen atoms at an internucleotide phosphorodiester backbone linkage. Like a natural phosphodiester RNA backbone linkage, a PS2-modified backbone linkage is achiral at phosphorus. PS2-RNAs are highly stable to nucleases and several in vitro assays have demonstrated their biological activity. For example, PS2-RNAs silenced mRNA in vitro and bound to protein targets in the form of PS2-aptamers (thioaptamers)...
September 18, 2017: Current Protocols in Nucleic Acid Chemistry
https://www.readbyqxmd.com/read/28921441/purification-and-autoactivation-method-for-recombinant-coagulation-factor-vii
#5
Vladimir Granovski, Marcela C C Freitas, Mario Soares Abreu-Neto, Dimas T Covas
Recombinant coagulation factor VII is a very important and complex protein employed for treatment of hemophiliac patients (hemophilia A/B) who develop inhibitors antibodies to conventional treatments (FVIII and FIX). The rFVII is a glycosylated molecule and circulates in plasma as zymogen of 50 kDa. When activated the molecule is cleaved to 20-30 kDa and has a half-life of about 3 h, needing to be processed fast and efficiently until freeze-drying. Here, we describe a very simple and fast purification sequence for rFVII using affinity FVII Select resin and a dialysis system that can be easily scaled up...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28921440/microplate-based-method-for-high-throughput-screening-hts-of-chromatographic-conditions-studies-for-recombinant-protein-purification
#6
Rimenys J Carvalho, Thayana A Cruz
High-throughput screening (HTS) systems have emerged as important tools to provide fast and low cost evaluation of several conditions at once since it requires small quantities of material and sample volumes. These characteristics are extremely valuable for experiments with large number of variables enabling the application of design of experiments (DoE) strategies or simple experimental planning approaches. Once, the capacity of HTS systems to mimic chromatographic purification steps was established, several studies were performed successfully including scale down purification...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28921439/purification-method-for-recombinant-hg-csf-by-affinity-chromatography
#7
Bruna Samham Archangelo, Elisa Maria de Sousa Russo
The human granulocytic colony-stimulating factor (hG-CSF) acts mainly by promoting the maturation of granulocytes and stimulating their phagocytic and chemotactic activity. It has been used in the treatment of many diseases, in particular in neutropenic conditions. Here, we describe the purification process of the recombinant protein hG-CSF expressed in Pichia pastoris. The protein purification proved to be efficient using the nickel affinity chromatography method described in this chapter.
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28921438/purification-methods-for-recombinant-factor-viii-expressed-in-human-liver-sk-hep-cells
#8
Vladimir Granovski, Mario Soares Abreu-Neto, Dimas Tadeu Covas
Coagulation factor VIII is one of the largest proteins attempted to be expressed in recombinant form. A very complex and labile protein which has a very short half-live and need a fast and efficient purification chain. Here, we describe a simple purification sequence using multimodal Capto MMC, affinity FVIII select and ion exchange SP-Fastflow chromatography steps without subjecting the target molecule to mechanical and temperature stress, separating impurities from rFVIII using net charge, hydrophobicity, and affinity of the molecules...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28921437/expression-of-glycosylated-proteins-in-bacterial-system-and-purification-by-affinity-chromatography
#9
Carlos Alexandre Breyer, Marcos Antonio de Oliveira, Adalberto Pessoa
The bacterial expression of glycoproteins has experienced significant progress in recent years, particularly in regard to the production of conjugate vaccines against pathogens. In this case, a protein carrier conjugated with glycosides is used to produce intense stimulation of the immune system. Glycoconjugate vaccines account for 35% of the global vaccine market, and consequently, several biotechnological companies have developed products for the purification of glycosylated proteins to attain homogeneity...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28921434/bioreactor-based-production-of-glycoproteins-in-plant-cell-suspension-cultures
#10
Tanja Holland, Johannes Felix Buyel
Recombinant glycoproteins such as monoclonal antibodies have a major impact on modern healthcare systems, e.g., as the active pharmaceutical ingredients in anticancer drugs. A specific glycan profile is often necessary to achieve certain desirable activities, such as the effector functions of an antibody, receptor binding or a sufficient serum half-life. However, many expression systems produce glycan profiles that differ substantially from the preferred form (usually the form found in humans) or produce a diverse array of glycans with a range of in vivo activities, thus necessitating laborious and costly separation and purification processes...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28921431/cell-free-production-of-protein-biologics-within-24-h
#11
Challise J Sullivan, Erik D Pendleton, John Dresios
Protein biologics have emerged as a safe and effective group of drug products that can be used in a variety of medical disorders and clinical settings, including treatment of orphan diseases, personalized medicine, and point-of-care applications. However, the full potential of protein biologics for such applications will not be realized until there are methods available for rapid and cost-effective production of small scale products for individual needs. Here, we describe a modular and scalable method for rapid and adaptable production of protein-based medical products at small doses...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28921426/production-of-full-length-antibody-by-pichia-pastoris
#12
Adam Nylen, Ming-Tang Chen
The methylotrophic yeast Pichia pastoris has become an increasingly popular host for recombinant protein expression in recent times. MRL pioneered a glycoengineered humanized P. pastoris expression system that could produce glycoproteins with glycosylation profiles similar to mammalian systems. Therapeutic glycoproteins produced by the humanized P. pastoris platform have shown comparable folding, stability, and in vitro and in vivo efficacies in preclinical models to their counterparts produced from the CHO cells...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28921304/expression-and-purification-of-an-fgf9-fusion-protein-in-e-coli-and-the-effects-of-the-fgf9-subfamily-on-human-hepatocellular-carcinoma-cell-proliferation-and-migration
#13
Shen Wang, Haipeng Lin, Tiantian Zhao, Sisi Huang, David G Fernig, Nuo Xu, Fenfang Wu, Mi Zhou, Chao Jiang, Haishan Tian
Fibroblast growth factor (FGF) 9 has oncogenic activity and plays an important role in the development of ovarian, lung, prostate, and gastric cancers. In the present study, with the aim of reducing the cost of utilizing growth factors in cancer research, a simple and efficient method for the preparation of recombinant human (rh)FGF9 in Escherichia coli was established. The rhFGF9 fusion protein (6 × His-TEV-rhFGF9) and the native protein released by tobacco etch virus (TEV) protease were obtained using a Ni-NTA system, with > 95% purity...
September 18, 2017: Applied Microbiology and Biotechnology
https://www.readbyqxmd.com/read/28920671/development-of-a-multifunctional-nanobiointerface-based-on-self-assembled-fusion-protein-rsbpa-zz-for-blood-cell-enrichment-and-phenotyping
#14
Mario Rothbauer, Martin Frauenlob, Karoline Gutkas, Michael B Fischer, Eva Kathrin Sinner, Seta Küpcü, Peter Ertl
We present a multifunctional nanobiointerface for blood cell capture and phenotyping applications that features both excellent antifouling properties and high antibody activity. Multifunctionality is accomplished by modifying polymeric materials using self-assembled S-layer fusion-protein rSbpA/ZZ to immobilize high density antibodies at the two protein A binding sites of the rSbpA/ZZ nanolattice structure. Controlled orientation and alignment of the antibodies reduced antibody consumption 100-fold and increased cell capture efficiency 4-fold over standard methodologies...
September 18, 2017: ACS Applied Materials & Interfaces
https://www.readbyqxmd.com/read/28920398/-fermentation-purification-and-immunogenicity-evaluation-of-hepatitis-e-virus-like-particles-expressed-in-hansenula-polymorpha
#15
Caixia Su, Li Li, Zhenji Jin, Xudong Han, Ping Zhao, Lin Wang, Chunhu Jiang, Yueli Wang, Wenwen Wang, Deqi Xu, Naishuo Zhu
To develop a new recombinant hepatitis E vaccine, we used Hansenula polymorpha expression system to express recombinant hepatitis E virus-like particles (HEV VLPs), to construct a recombinant engineered strain HP/HEV2.3. The fermentation conditions and purification process were studied next. The first working seed lots were fermented in liquid culture, and the fermentation products were collected, then crushed, clarified, purified by ultrafiltration, silica gel adsorbed and desorbed, concentrated by ultrafiltration, purified by liquid chromatography and sterilized by filtration...
April 25, 2017: Sheng Wu Gong Cheng Xue Bao, Chinese Journal of Biotechnology
https://www.readbyqxmd.com/read/28920164/purification-and-characterization-of-a-naringinase-from-cryptococcus-albidus
#16
Nataliya Borzova, Olena Gudzenko, Lyudmila Varbanets
Naringinase which was extracted from the fermented broth of Cryptococcus albidus was purified about 42-folds with yield 0.7% by sulfate fractionation and chromatography on Toyopearl HW-60, Fractogel DEAE-650-s, and Sepharose 6B columns. Molecular weight of protein determined by gel filtration and SDS-PAGE was 50 kDa. Naringinase of C. albidus includes high content of the dicarbonic and hydrophobic amino acids. Enzyme contains also carbohydrate component, represented by mannose, galactose, rhamnose, ribose, arabinose, xylose, and glucose...
September 18, 2017: Applied Biochemistry and Biotechnology
https://www.readbyqxmd.com/read/28919250/purification-and-characterization-of-a-cellulase-free-thermostable-endo-xylanase-from-streptomyces-griseorubens-lh-3-and-its-use-in-biobleaching-on-eucalyptus-kraft-pulp
#17
Hao Wu, Xianbo Cheng, Yongfeng Zhu, Wei Zeng, Guiguang Chen, Zhiqun Liang
Xylanase is an important enzyme involved in degrading xylan. In this study, an extracellular cellulase-free, thermostable endo-xylanase which was produced by Streptomyces griseorubens LH-3 with bagasse semi-cellulose as a carbon source was purified and characterized. The xylanase was purified 4-fold with a recovery yield of 21.6% by precipitation with 25-55% (NH4)2SO4, Mono Q ion exchange chromatography and sephacryl S-200 HR gel filtration chromatography. It appeared as a monomeric protein on SDS-PAGE gel and had an apparent molecular weight of 45...
September 14, 2017: Journal of Bioscience and Bioengineering
https://www.readbyqxmd.com/read/28918196/expression-and-purification-of-human-and-saccharomyces-cerevisiae-equilibrative-nucleoside-transporters
#18
Rebba C Boswell-Casteel, Jennifer M Johnson, Zygy Roe-Žurž, Kelli D Duggan, Hannah Schmitz, Franklin A Hays
Nucleosides play an essential role in the physiology of eukaryotes by acting as metabolic precursors in de novo nucleic acid synthesis and energy metabolism. Nucleosides also act as ligands for purinergic receptors. Equilibrative nucleoside transporters (ENTs) are polytopic integral membrane proteins that aid in regulating plasmalemmal flux of purine and pyrimidine nucleosides and nucleobases. ENTs exhibit broad substrate selectivity across different isoforms and utilize diverse mechanisms to drive substrate flux across membranes...
September 13, 2017: Protein Expression and Purification
https://www.readbyqxmd.com/read/28917863/purification-characterization-and-biological-effect-of-reversing-the-kidney-yang-deficiency-of-polysaccharides-from-semen-cuscutae
#19
Song Yang, Xinfang Xu, Hefang Xu, Shuya Xu, Qinghua Lin, Zhe Jia, Ting Han, Hui Zhang, Ying Zhang, Huan Liu, Yan Gao, Xiangri Li
Semen cuscutae is a well-known Chinese medicine which has been used to nourish kidney. It is the first study to demonstrate that the polysaccharides from semen cuscutae showed significant activity of nourishing kidney-yang by increasing the levels of testosterone and estradiol, decreasing the level of blood urea nitrogen, improving immune function, possessing antioxidant effect. Three homogeneous polysaccharides were obtained by DEAE-cellulose and Sephacryl S-400 which were named as C-7WR1, C-7WR2 and C-7WR3 with average molecular weight of 7...
November 1, 2017: Carbohydrate Polymers
https://www.readbyqxmd.com/read/28915418/purification-identification-and-molecular-mechanism-of-two-dipeptidyl-peptidase-iv-dpp-iv-inhibitory-peptides-from-antarctic-krill-euphausia-superba-protein-hydrolysate
#20
Wei Ji, Chaohua Zhang, Hongwu Ji
Dipeptidyl peptidase IV (DPP-IV) played an important role in blood glucose regulation. Inhibition of DPP-IV may improve glycemic control in diabetics by preventing the rapid breakdown of incretin hormones and prolonging their physiological action. In this study, Antarctic krill (Euphausia superba) protein was hydrolyzed using animal proteolytic enzymes. The hydrolysate was purified sequentially by ultrafiltration, gel filtration chromatography and reversed phase high-performance liquid chromatography (RP-HPLC)...
September 5, 2017: Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences
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