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51.
Identifying the membrane proteome of HIV-1 latently infected cells 总被引:11,自引:0,他引:11
Berro R de la Fuente C Klase Z Kehn K Parvin L Pumfery A Agbottah E Vertes A Nekhai S Kashanchi F 《The Journal of biological chemistry》2007,282(11):8207-8218
Profiling integral plasma membrane proteins is of particular importance for the identification of new biomarkers for diagnosis and for drug development. We report in this study the identification of surface markers by performing comparative proteomics of established human immunodeficiency virus-1 (HIV-1) latent cell models and parental cell lines. To this end we isolated integral membrane proteins using a biotin-directed affinity purification method. Isolated proteins were separated by two-dimensional gel electrophoresis and identified by matrix-assisted laser desorption/ionization-time-of-flight (MALDI-TOF) after in gel digestion. Seventeen different proteins were found to vary on the surface of T-cells due to HIV-1 infection. Of these proteins, 47% were integral membrane proteins, and 18% were membrane-associated. Through the use of complementary techniques such as Western blotting and fluorescent staining, we confirmed the differential expression of some of the proteins identified by MALDI-TOF including Bruton's tyrosine kinase and X-linked inhibitor of apoptosis. Finally, using phosphatidylinositol 3-kinase inhibitors and flavopiridol to inhibit Bruton's tyrosine kinase localization at the membrane and X-linked inhibitor of apoptosis protein expression, respectively, we showed that HIV-1 latently infected cells are more sensitive to these drugs than uninfected cells. This suggests that HIV-1 latently infected cells may be targeted with drugs that alter several pathways that are essential for the establishment and maintenance of latency. 相似文献
52.
Storozhenko S Navarrete O Ravanel S De Brouwer V Chaerle P Zhang GF Bastien O Lambert W Rébeillé F Van Der Straeten D 《The Journal of biological chemistry》2007,282(14):10749-10761
In plants, 6-hydroxymethyl-7,8-dihydropterin pyrophosphokinase/7,8-dihydropteroate synthase (mitHPPK/DHPS) is a bifunctional mitochondrial enzyme, which catalyzes the first two consecutive steps of tetrahydrofolate biosynthesis. Mining the Arabidopsis genome data base has revealed a second gene encoding a protein that lacks a potential transit peptide, suggesting a cytosolic localization of the isoenzyme (cytHPPK/DHPS). When the N-terminal part of the cytHPPK/DHPS was fused to green fluorescent protein, the fusion protein appeared only in the cytosol, confirming the above prediction. Functionality of cytHPPK/DHPS was addressed by two parallel approaches: first, the cytHPPK/DHPS was able to rescue yeast mutants lacking corresponding activities; second, recombinant cytHPPK/DHPS expressed and purified from Escherichia coli displayed both HPPK and DHPS activities in vitro. In contrast to mitHPPK/DHPS, which was ubiquitously expressed, the cytHPPK/DHPS gene was exclusively expressed in reproductive tissue, more precisely in developing seeds as revealed by histochemical analysis of a transgenic cytHPPK/DHPS promoter-GUS line. In addition, it was observed that expression of cytHPPK/DHPS mRNA was induced by salt stress, suggesting a potential role of the enzyme in stress response. This was supported by the phenotype of a T-DNA insertion mutant in the cytHPPK/DHPS gene, resulting in lower germination rates as compared with the wild-type upon application of oxidative and osmotic stress. 相似文献
53.
Chernyavsky AI Arredondo J Kitajima Y Sato-Nagai M Grando SA 《The Journal of biological chemistry》2007,282(18):13804-13812
Although it is accepted that pemphigus antibody binding to keratinocytes (KCs) evokes an array of intracellular biochemical events resulting in cell detachment and death, the triggering events remain obscure. It has been postulated that the binding of pemphigus vulgaris IgG (PVIgG) to KCs induces "desmosomal" signaling. Because in contrast to integrins and classical cadherins, desmoglein (Dsg) molecules are not known to elicit intracellular signaling, and because PV patients also produce non-Dsg autoantibodies, we investigated the roles of both Dsg and non-desmoglein PV antigens. The time course studies of KCs treated with PVIgG demonstrated that the activity of Src peaked at 30 min, EGF receptor kinase (EGFRK) at 60 min, and p38 MAPK at 240 min. The Src inhibitor PP2 decreased EGFRK and p38 activities by approximately 45 and 30%, respectively, indicating that in addition to Src, PVIgG evokes other triggering events. The shrinkage of KCs (cell volume reduction) became significant at 120 min, keratin aggregation at 240 min, and an increase of TUNEL positivity at 360 min. Pretreatment of KCs with PP2 blocked PVIgG-dependent cell shrinkage and keratin aggregation by approximately 50% and TUNEL positivity by approximately 25%. The p38 MAPK inhibitor PD169316 inhibited these effects by approximately 15, 20, and 70%, respectively. Transfection of KCs with small interfering RNAs that silenced expression of Dsg1 and/or Dsg3 proteins, blocked approximately 50% of p38 MAPK activity but did not significantly alter the PVIgG-dependent rise in Src and EGFRK activities. These results indicate that activation of p38 MAPK is a late signaling step associated with collapse of the cytoskeleton and disassembly of desmosomes caused by upstream events involving Src and EGFRK. Therefore, the early acantholytic events are triggered by non-Dsg antibodies. 相似文献
54.
55.
Typical physico-chemical studies of metal binding proteins are usually aimed at determination of the metal binding constant
K for a native protein (K
n), while the significance of the K value for the thermally denatured protein (K
u) is usually underestimated. Meanwhile, metal binding induced shift of thermal denaturation transition of a single site metal
binding protein is defined by K
n to K
u ratio, implying that knowledge of both K values is required for full characterization of the system. In the present work, the most universal approach to the studies
of single site metal binding proteins, namely construction of a protein “phase diagram” in coordinates of free metal ion concentration
– temperature, is considered in detail. The detailed algorithm of construction of the phase diagrams along with underlying
mathematic procedures developed here may be of use for studies of other simple protein-target type systems, where target represents
low molecular weight ligand. Analysis of the simplest protein-ligand system reveals that thermodynamic properties of apo-protein
dictate the maximal possible increase of its affinity to any simple ligand upon thermal denaturation of the protein. Experimental
and general problems coupled with the use of the phase diagrams are discussed. 相似文献
56.
57.
Sergei Shalygin I‐Shuo Huang Elle H. Allen JoAnn M. Burkholder Paul V. Zimba 《Journal of phycology》2019,55(3):509-520
Pleurocapsales are one of the least understood groups of cyanobacteria in terms of molecular systematics and biochemistry. Considering the high number of cryptic taxa within the Synechococcales and Oscillatoriales, it is likely that such taxa also occur in the Pleurocapsales. The new genus described in our research is the first known pleurocapsalean cryptic taxon. It produces off‐flavor and a large number of bioactive metabolites (n = 38) some of which can be toxic including four known microcystins. Using a polyphasic approach, we propose the establishment of the genus Odorella with the new species O. benthonica from material originally isolated from the California Aqueduct near Los Angeles. 相似文献
58.
Taras Antal Ivan Konyukhov Alena Volgusheva Tatyana Plyusnina Sergei Khruschev Galina Kukarskikh Sergey Goryachev Andrey Rubin 《Physiologia plantarum》2019,165(3):476-486
The development of high‐performance photobioreactors equipped with automatic systems for non‐invasive real‐time monitoring of cultivation conditions and photosynthetic parameters is a challenge in algae biotechnology. Therefore, we developed a chlorophyll (Chl) fluorescence measuring system for the online recording of the light‐induced fluorescence rise and the dark relaxation of the flash‐induced fluorescence yield (Qa? ? re‐oxidation kinetics) in photobioreactors. This system provides automatic measurements in a broad range of Chl concentrations at high frequency of gas‐tight sampling, and advanced data analysis. The performance of this new technique was tested on the green microalgae Chlamydomonas reinhardtii subjected to a sulfur deficiency stress and to long‐term dark anaerobic conditions. More than thousand fluorescence kinetic curves were recorded and analyzed during aerobic and anaerobic stages of incubation. Lifetime and amplitude values of kinetic components were determined, and their dynamics plotted on heatmaps. Out of these data, stress‐sensitive kinetic parameters were specified. This implemented apparatus can therefore be useful for the continuous real‐time monitoring of algal photosynthesis in photobioreactors. 相似文献
59.
Scribano Vittorio Simakov Sergei K. Finocchiaro Claudio Correale Alessandra Scirè Salvatore 《Origins of life and evolution of the biosphere》2019,49(1-2):19-47
Origins of Life and Evolution of Biospheres - Pyrite and organic matter closely coexist in some hydrothermally-altered gabbroic xenoliths from the Hyblean Plateau, Sicily. The representative sample... 相似文献
60.
Permyakova Natalya V. Sidorchuk Yury V. Marenkova Tatyana V. Khozeeva Sofya A. Kuznetsov Vitaly V. Zagorskaya Alla A. Rozov Sergei M. Deineko Elena V. 《Molecular biology reports》2019,46(6):5735-5743
Molecular Biology Reports - Targeted genome editing using CRISPR/Cas9 is a promising technology successfully verified in various plant species; however, it has hardly been used in plant cell... 相似文献