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Daniel M. Strauss Scott Lute Zinaida Tebaykina Douglas D. Frey Cintia Ho Gregory S. Blank Kurt Brorson Qi Chen Bin Yang 《Biotechnology and bioengineering》2009,104(2):371-380
During production of therapeutic monoclonal antibodies (mAbs) in mammalian cell culture, it is important to ensure that viral impurities and potential viral contaminants will be removed during downstream purification. Anion exchange chromatography provides a high degree of virus removal from mAb feedstocks, but the mechanism by which this is achieved has not been characterized. In this work, we have investigated the binding of three viruses to Q sepharose fast flow (QSFF) resin to determine the degree to which electrostatic interactions are responsible for viral clearance by this process. We first used a chromatofocusing technique to determine the isoelectric points of the viruses and established that they are negatively charged under standard QSFF conditions. We then determined that virus removal by this chromatography resin is strongly disrupted by the presence of high salt concentrations or by the absence of the positively charged Q ligand, indicating that binding of the virus to the resin is primarily due to electrostatic forces, and that any non‐electrostatic interactions which may be present are not sufficient to provide virus removal. Finally, we determined the binding profile of a virus in a QSFF column after a viral clearance process. These data indicate that virus particles generally behave similarly to proteins, but they also illustrate the high degree of performance necessary to achieve several logs of virus reduction. Overall, this mechanistic understanding of an important viral clearance process provides the foundation for the development of science‐based process validation strategies to ensure viral safety of biotechnology products. Biotechnol. Bioeng. 2009; 104: 371–380 © 2009 Wiley Periodicals, Inc. 相似文献
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Michael Lauck Sergey V. Alkhovsky Yīmíng Bào Adam L. Bailey Zinaida V. Shevtsova Alexey M. Shchetinin Tatyana V. Vishnevskaya Matthew G. Lackemeyer Elena Postnikova Steven Mazur Jiro Wada Sheli R. Radoshitzky Thomas C. Friedrich Boris A. Lapin Petr G. Deriabin Peter B. Jahrling Tony L. Goldberg David H. O'Connor Jens H. Kuhn 《Journal of virology》2015,89(15):8082-8087
Simian hemorrhagic fever (SHF) is lethal for macaques. Based on clinical presentation and serological diagnosis, all reported SHF outbreaks were thought to be caused by different strains of the same virus, simian hemorrhagic fever virus (SHFV; Arteriviridae). Here we show that the SHF outbreaks in Sukhumi in 1964 and in Alamogordo in 1989 were caused not by SHFV but by two novel divergent arteriviruses. Our results indicate that multiple divergent simian arteriviruses can cause SHF. 相似文献
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Zinaida Doubeikovskaia Anne Aries Pierre Jeannesson Francois Morle Alexandre Doubeikovski 《Protein expression and purification》2001,23(3):426-431
GATA-1 is a key regulator of terminal erythroid differentiation in mammals and birds. The structural and biochemical studies of human GATA-1 (hGATA-1) are limited by the difficulty of its purification in a sufficient amount. Here we describe the procedure for obtaining pure bacterial recombinant hGATA-1 in an active functional state. We demonstrate that this protein may be successfully used for preparing an affinity column, producing GATA-1-specific rabbit polyclonal antibodies, and studying DNA-protein and protein-protein interactions. 相似文献
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Hall AH Wan J Spesock A Sergueeva Z Shaw BR Alexander KA 《Nucleic acids research》2006,34(9):2773-2781
In RNA interference (RNAi), double-stranded short interfering RNA (ds-siRNA) inhibits expression from complementary mRNAs. Recently, it was demonstrated that short, single-stranded antisense RNA (ss-siRNA) can also induce RNAi. While ss-siRNA may offer several advantages in both clinical and research applications, its overall poor activity compared with ds-siRNA has prevented its widespread use. In contrast to the poor gene silencing activity of native ss-siRNA, we found that the silencing activity of boranophosphate-modified ss-siRNA is comparable with that of unmodified ds-siRNA. Boranophosphate ss-siRNA has excellent maximum silencing activity and is highly effective at low concentrations. The silencing activity of boranophosphate ss-siRNA is also durable, with significant silencing up to 1 week after transfection. Thus, we have demonstrated that boranophosphate-modified ss-siRNA can silence gene expression as well as native ds-siRNA, suggesting that boranophosphate-modified ss-siRNAs should be investigated as a potential new class of therapeutic agents. 相似文献
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Rafayel A. Azizyan Adriana Garro Zinaida Radkova Alexey Anikeenko Anastasia Bakulina Christian Dumas Andrey V. Kajava 《Journal of molecular biology》2018,430(20):3835-3846
In many disease-related and functional amyloids, the amyloid-forming regions of proteins are flanked by globular domains. When located in close vicinity of the amyloid regions along the chain, the globular domains can prevent the formation of amyloids because of the steric repulsion. Experimental tests of this effect are few in number and non-systematic, and their interpretation is hampered by polymorphism of amyloid structures. In this situation, modeling approaches that use such a clear-cut criterion as the steric tension can give us highly trustworthy results. In this work, we evaluated this steric effect by using molecular modeling and dynamics. As an example, we tested hybrid proteins containing an amyloid-forming fragment of Aβ peptide (17–42) linked to one or two globular domains of GFP. Searching for the shortest possible linker, we constructed models with pseudo-helical arrangements of the densely packed GFPs around the Aβ amyloid core. The molecular modeling showed that linkers of 7 and more residues allow fibrillogenesis of the Aβ-peptide flanked by GFP on one side and 18 and more residues when Aβ-peptide is flanked by GFPs on both sides. Furthermore, we were able to establish a more general relationship between the size of the globular domains and the length of the linkers by using analytical expressions and rigid body simulations. Our results will find use in planning and interpretation of experiments, improvement of the prediction of amyloidogenic regions in proteins, and design of new functional amyloids carrying globular domains. 相似文献
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Dauphinee SM Voelcker V Tebaykina Z Wong F Karsan A 《American journal of physiology. Heart and circulatory physiology》2011,301(6):H2246-H2253
The innate immune recognition of bacterial lipopolysaccharide (LPS) is mediated by Toll-like receptor 4 (TLR4) and results in activation of proinflammatory signaling including NF-κB and MAPK pathways. Heterotrimeric G proteins have been previously implicated in LPS signaling in macrophages and monocytes. In the present study, we show that pertussis toxin sensitive heterotrimeric G proteins (Gα(i/o)) are involved in the activation of MAPK and Akt downstream of TLR2, TLR3, and TLR4 in endothelial cells. Gα(i/o) are also required for full activation of interferon signaling downstream of TLR3 and TLR4 but are not required for the activation of NF-κB. We find that Gα(i/o)-mediated activation of the MAPK is independent of the canonical MyD88, interleukin-1 receptor-associated kinase, and tumor necrosis factor receptor-associated factor 6 signaling cascade in LPS-stimulated cells. Taken together, the data presented here suggest that heterotrimeric G proteins are widely involved in TLR pathways along a signaling cascade that is distinct from MyD88-TRAF6. 相似文献
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Nadezda V. Amaegberi Galina N. Semenkova Zinaida B. Kvacheva Alexandra G. Lisovskaya Serge V. Pinchuk Oleg I. Shadyro 《Cell biochemistry and function》2019,37(4):281-289
2‐Hexadecenal (2HD) formation in the organism occurs via irreversible enzymatic degradation of sphingosine‐1‐phosphate or nonenzymatic γ‐, UV‐, or HOCl‐induced destruction of a number of sphingolipids including S1P. The current research focuses on the study of 2HD effects on C6 glioma cells growth. The results obtained show that 2HD causes a dose‐dependent decrease in proliferative and mitotic indices. The change in the mitotic index is due to the redistribution of cells in the different phases of mitosis. These processes are accompanied by cytoskeleton rearrangement and changes in cell morphology, which are expressed in F‐actin redistribution, change in the number and type of filopodia and fibrils, leading to cell shape changes, decrease in intercellular contacts and monolayer rarefaction. Cells treatment with 2HD leads to apoptosis induction and signalling pathways modification, including activation of JNK, p38, and ERK1/2 MAPK but not PI3K. The effects observed are not related to the cytotoxicity of 2HD. Significance of the study: 2HD—an unsaturated aldehyde, which level can rise under conditions of oxidative stress as a result of nonenzymatic sphingolipids' destruction. The mechanisms of 2HD action on various cell types have not been sufficiently studied. Therefore, the study on functional role of this aldehyde in different cell types that may be its target is relevant. This study demonstrated that 2HD inhibits growth of C6 glioma cells due to modification of intracellular processes of signal transduction, cytoskeleton rearrangement, change in the mitotic regimen and apoptosis induction. 相似文献
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Jenkins R Bobyleva Z Goldberg D Gask L Zacroeva AG Potasheva A Krasnov V McDaid D 《Mental health in family medicine》2009,6(1):29-36
Introduction Mental disorders occur as frequently in Russia as elsewhere, but the common mental disorders, especially depression, have gone largely unrecognised and undiagnosed by policlinic staff and area doctors.Methods This paper describes the impact and sustainability of a multi-component programme to facilitate the integration of mental health into primary care, by situation appraisal, policy dialogue, development of educational materials, provision of a training programme and the publication of standards and good practice guidelines to improve the primary care of mental disorders in the Sverdlovsk region of the Russian Federation.Results The multi-component programme has resulted in sustainable training about common mental disorders, not only of family doctors but also of other cadres and levels of professionals, and it has been well integrated with Sverdlovsk's overall programme of health sector reforms.Conclusion It is possible to facilitate the sustainable integration of mental health into primary care within the Russian context. While careful adaptation will be needed, the approach adopted here may also hold useful lessons for policy makers seeking to integrate mental health within primary care in other contexts and settings. 相似文献