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51.
Rabah Gahoual Alicia Burr Jean-Marc Busnel Lauriane Kuhn Phillipe Hammann Alain Beck Yannis-Nicolas Fran?ois Emmanuelle Leize-Wagner 《MABS-AUSTIN》2013,5(3):479-490
Monoclonal antibodies (mAbs) are highly complex proteins that display a wide range of microheterogeneity that requires multiple analytical methods for full structure assessment and quality control. As a consequence, the characterization of mAbs on different levels is particularly product - and time - consuming. This work presents the characterization of trastuzumab sequence using sheathless capillary electrophoresis (referred as CESI) – tandem mass spectrometry (CESI-MS/MS). Using this bottom-up proteomic-like approach, CESI-MS/MS provided 100% sequence coverage for both heavy and light chain via peptide fragment fingerprinting (PFF) identification. The result was accomplished in a single shot, corresponding to the analysis of 100 fmoles of digest. The same analysis also enabled precise characterization of the post-translational hot spots of trastuzumab, used as a representative widely marketed therapeutic mAb, including the structural confirmation of the five major N-glycoforms. 相似文献
52.
Pablo A. Imbach Bruno Locatelli Luis G. Molina Philippe Ciais Paul W. Leadley 《Ecology and evolution》2013,3(9):2917-2932
Climate change is a threat to biodiversity, and adaptation measures should be considered in biodiversity conservation planning. Protected areas (PA) are expected to be impacted by climate change and improving their connectivity with biological corridors (BC) has been proposed as a potential adaptation measure, although assessing its effectiveness remains a challenge. In Mesoamerica, efforts to preserve the biodiversity have led to the creation of a regional network of PA and, more recently, BC. This study evaluates the role of BC for facilitating plant dispersal between PA under climate change in Mesoamerica. A spatially explicit dynamic model (cellular automaton) was developed to simulate species dispersal under different climate and conservation policy scenarios. Plant functional types (PFT) were defined based on a range of dispersal rates and vegetation types to represent the diversity of species in the region. The impacts of climate change on PA and the role of BC for dispersal were assessed spatially. Results show that most impacted PA are those with low altitudinal range in hot, dry, or high latitude areas. PA with low altitudinal range in high cool areas benefit the most from corridors. The most important corridors cover larger areas and have high altitude gradients. Only the fastest PFT can keep up with the expected change in climate and benefit from corridors for dispersal. We conclude that the spatial assessment of the vulnerability of PA and the role of corridors in facilitating dispersal can help conservation planning under a changing climate. 相似文献
53.
Lauton-Santos S Guatimosim S Castro CH Oliveira FA Almeida AP Dias-Peixoto MF Gomes MA Pessoa P Pesquero JL Pesquero JB Bader M Cruz JS 《Life sciences》2007,81(10):814-822
The kinins have an important role in control of the cardiovascular system. They have been associated with protective effects in the heart tissue. Kinins act through stimulation of two 7-transmembrane G protein-coupled receptors, denoted B(1) and B(2) receptors. However, the physiological relevance of B(1) receptor in the heart has not been clearly established. Using B(1) kinin receptor gene knock-out mice we tested the hypothesis that the B(1) receptor plays an important role in the control of baseline cardiac function. We examined the functional aspects of the intact heart and also in the isolated cardiomyocytes to study intracellular Ca(2+) cycling by using confocal microscopy and whole-cell voltage clamp techniques. We measured heart rate, diastolic and systolic tension, contraction and relaxation rates and, coronary perfusion pressure. Whole-cell voltage clamp was performed to measure L-type Ca(2+) current (I(Ca,L)). The hearts from B(1)(-/-) mice showed smaller systolic tension. The average values for WT and B(1)(-/-) mice were 2.6+/-0.04 g vs. 1.6+/-0.08 g, respectively. This result can be explained, at least in part, by the decrease in the Ca(2+) transient (3.1+/-0.06 vs. 3.4+/-0.09 for B(1)(-/-) and WT, respectively). There was an increase in I(Ca,L) at depolarized membrane potentials. Interestingly, the inactivation kinetics of I(Ca,L) was statistically different between the groups. The coronary perfusion pressure was higher in the hearts from B(1)(-/-) mice indicating an increase in coronary resistance. This result can be explained by the significant reduction of eNOS (NOS-3) expression in the aorta of B(1)(-/-) mice. Collectively, our results demonstrate that B(1) receptor exerts a fundamental role in the mammalian cardiac function. 相似文献
54.
Thyroid hormone excess rather than thyrotropin deficiency induces osteoporosis in hyperthyroidism 总被引:6,自引:0,他引:6
Bassett JH O'Shea PJ Sriskantharajah S Rabier B Boyde A Howell PG Weiss RE Roux JP Malaval L Clement-Lacroix P Samarut J Chassande O Williams GR 《Molecular endocrinology (Baltimore, Md.)》2007,21(5):1095-1107
Thyrotoxicosis is an important but under recognized cause of osteoporosis. Recently, TSH deficiency, rather than thyroid hormone excess, has been suggested as the underlying cause. To investigate the molecular mechanism of osteoporosis in thyroid disease, we characterized the skeleton in mice lacking either thyroid hormone receptor alpha or beta (TRalpha(0/0), TRbeta-/-). Remarkably, in the presence of normal circulating thyroid hormone and TSH concentrations, adult TRalpha(0/0) mice had osteosclerosis accompanied by reduced osteoclastic bone resorption, whereas juveniles had delayed endochondral ossification with reduced bone mineral deposition. By contrast, adult TRbeta-/- mice with elevated TSH and thyroid hormone levels were osteoporotic with evidence of increased bone resorption, whereas juveniles had advanced ossification with increased bone mineral deposition. Analysis of T3 target gene expression revealed skeletal hypothyroidism in TRalpha(0/0) mice, but skeletal thyrotoxicosis in TRbeta-/- mice. These studies demonstrate that bone loss in thyrotoxicosis is independent of circulating TSH levels and mediated predominantly by TRalpha, thus identifying TRalpha as a novel drug target in the prevention and treatment of osteoporosis. 相似文献
55.
Ann R. Stavert Marielle Saunois Josep G. Canadell Benjamin Poulter Robert B. Jackson Pierre Regnier Ronny Lauerwald Peter A. Raymond George H. Allen Prabir K. Patra Peter Bergamaschi Phillipe Bousquet Naveen Chandra Philippe Ciais Adrian Gustafson Misa Ishizawa Akihiko Ito Thomas Kleinen Shamil Maksyutov Joe McNorton Joe R. Melton Jurek Müller Yosuke Niwa Shushi Peng William J. Riley Arjo Segers Hanqin Tian Aki Tsuruta Yi Yin Zhen Zhang Bo Zheng Qianlai Zhuang 《Global Change Biology》2022,28(1):182-200
The ongoing development of the Global Carbon Project (GCP) global methane (CH4) budget shows a continuation of increasing CH4 emissions and CH4 accumulation in the atmosphere during 2000–2017. Here, we decompose the global budget into 19 regions (18 land and 1 oceanic) and five key source sectors to spatially attribute the observed global trends. A comparison of top-down (TD) (atmospheric and transport model-based) and bottom-up (BU) (inventory- and process model-based) CH4 emission estimates demonstrates robust temporal trends with CH4 emissions increasing in 16 of the 19 regions. Five regions—China, Southeast Asia, USA, South Asia, and Brazil—account for >40% of the global total emissions (their anthropogenic and natural sources together totaling >270 Tg CH4 yr?1 in 2008–2017). Two of these regions, China and South Asia, emit predominantly anthropogenic emissions (>75%) and together emit more than 25% of global anthropogenic emissions. China and the Middle East show the largest increases in total emission rates over the 2000 to 2017 period with regional emissions increasing by >20%. In contrast, Europe and Korea and Japan show a steady decline in CH4 emission rates, with total emissions decreasing by ~10% between 2000 and 2017. Coal mining, waste (predominantly solid waste disposal) and livestock (especially enteric fermentation) are dominant drivers of observed emissions increases while declines appear driven by a combination of waste and fossil emission reductions. As such, together these sectors present the greatest risks of further increasing the atmospheric CH4 burden and the greatest opportunities for greenhouse gas abatement. 相似文献
56.
Rafael Molina Pilar Redondo Blanca López-Méndez Maider Villate Nekane Merino Francisco J. Blanco Julien Valton Silvestre Grizot Phillipe Duchateau Jesús Prieto Guillermo Montoya 《The Journal of biological chemistry》2015,290(48):28727-28736
Homing endonucleases recognize and generate a DNA double-strand break, which has been used to promote gene targeting. These enzymes recognize long DNA stretches; they are highly sequence-specific enzymes and display a very low frequency of cleavage even in complete genomes. Although a large number of homing endonucleases have been identified, the landscape of possible target sequences is still very limited to cover the complexity of the whole eukaryotic genome. Therefore, the finding and molecular analysis of homing endonucleases identified but not yet characterized may widen the landscape of possible target sequences. The previous characterization of protein-DNA interaction before the engineering of new homing endonucleases is essential for further enzyme modification. Here we report the crystal structure of I-CvuI in complex with its target DNA and with the target DNA of I-CreI, a homologue enzyme widely used in genome engineering. To characterize the enzyme cleavage mechanism, we have solved the I-CvuI DNA structures in the presence of non-catalytic (Ca2+) and catalytic ions (Mg2+). We have also analyzed the metal dependence of DNA cleavage using Mg2+ ions at different concentrations ranging from non-cleavable to cleavable concentrations obtained from in vitro cleavage experiments. The structure of I-CvuI homing endonuclease expands the current repertoire for engineering custom specificities, both by itself as a new scaffold alone and in hybrid constructs with other related homing endonucleases or other DNA-binding protein templates. 相似文献
57.
Christelle Mbondji-Wonje Viswanath Ragupathy Jiangqin Zhao Aubin Nanfack Sherwin Lee Judith Torimiro Phillipe Nyambi Indira K. Hewlett 《PloS one》2014,9(11)
Background
The use of CCR5 antagonists involves determination of HIV-1 tropism prior to initiation of treatment. HIV-1 tropism can be assessed either by phenotypic or genotypic methods. Genotypic methods are extensively used for tropism prediction. However, their validation in predicting tropism of viral isolates belonging to group M non-B subtypes remains challenging. In Cameroon, the genetic diversity of HIV-1 strains is the broadest reported worldwide. To facilitate the integration of CCR5 antagonists into clinical practice in this region, there is a need to evaluate the performance of genotypic methods for predicting tropism of highly diverse group M HIV-1 strains.Methods
Tropism of diverse HIV-1 strains isolated from PBMCs from Cameroon was determined using the GHOST cell assay. Prediction, based on V3 sequences from matched plasma samples, was determined using bioinformatics algorithms and rules based on position 11/25 and net charge applied independently or combined according to Delobel''s and Garrido''s rules. Performance of genotypic methods was evaluated by comparing prediction generated with tropism assigned by the phenotypic assay.Results
Specificity for predicting R5-tropic virus was high, ranging from 83.7% to 97.7% depending on the genotypic methods used. Sensitivity for X4-tropic viruses was fairly low, ranging from 33.3% to 50%. In our study, overall, genotypic methods were less able to accurately predict X4-tropic virus belonging to subtype CRF02_AG. In addition, it was found that of the methods we used the Garrido rule has the highest sensitivity rate of over 50% with a specificity of 93%.Conclusion
Our study demonstrated that overall, genotypic methods were less sensitive for accurate prediction of HIV-1 tropism in settings where diverse HIV-1 strains co-circulate. Our data suggest that further optimization of genotypic methods is needed and that larger studies to determine their utility for tropism prediction of diverse HIV-1 strains may be warranted. 相似文献58.
Odile Filhol Delphine Ciais Christian Lajaunie Peggy Charbonnier Nicolas Foveau Jean-Philippe Vert Yves Vandenbrouck 《PloS one》2012,7(10)
Chemically synthesized small interfering RNA (siRNA) is a widespread molecular tool used to knock down genes in mammalian cells. However, designing potent siRNA remains challenging. Among tools predicting siRNA efficacy, very few have been validated on endogenous targets in realistic experimental conditions. We previously described a tool to assist efficient siRNA design (DSIR, Designer of siRNA), which focuses on intrinsic features of the siRNA sequence. Here, we evaluated DSIR’s performance by systematically investigating the potency of the siRNA it designs to target ten cancer-related genes. mRNA knockdown was measured by quantitative RT-PCR in cell-based assays, revealing that over 60% of siRNA sequences designed by DSIR silenced their target genes by at least 70%. Silencing efficacy was sustained even when low siRNA concentrations were used. This systematic analysis revealed in particular that, for a subset of genes, the efficiency of siRNA constructs significantly increases when the sequence is located closer to the 5′-end of the target gene coding sequence, suggesting the distance to the 5′-end as a new feature for siRNA potency prediction. A new version of DSIR incorporating these new findings, as well as the list of validated siRNA against the tested cancer genes, has been made available on the web (http://biodev.extra.cea.fr/DSIR). 相似文献
59.
Recent studies have demonstrated that both the potency and breadth of the humoral anti-HIV-1 immune response in generating neutralizing antibodies (nAbs) against heterologous viruses are significantly enhanced after superinfection by discordant HIV-1 subtypes, suggesting that repeated exposure of the immune system to highly diverse HIV-1 antigens can significantly improve anti-HIV-1 immunity. Thus, we investigated whether sequential plasma from these subjects superinfected with discordant HIV-1 subtypes, who exhibit broad nAbs against heterologous viruses, also neutralize their discordant early autologous viruses with increasing potency. Comparing the neutralization capacities of sequential plasma obtained before and after superinfection of 4 subjects to those of matched plasma obtained from 4 singly infected control subjects, no difference in the increase in neutralization capacity was observed between the two groups (p = 0.328). Overall, a higher increase in neutralization over time was detected in the singly infected patients (mean change in IC(50) titer from first to last plasma sample: 183.4) compared to the superinfected study subjects (mean change in IC(50) titer from first to last plasma sample: 66.5). Analysis of the Breadth-Potency Scores confirmed that there was no significant difference in the increase in superinfected and singly infected study subjects (p = 0.234). These studies suggest that while superinfection by discordant subtypes induces antibodies with enhanced neutralizing breadth and potency against heterologous viruses, the potency to neutralize their autologous viruses is not better than those seen in singly infected patients. 相似文献
60.
Terrestrial biosphere models need better representation of vegetation phenology: results from the North American Carbon Program Site Synthesis 总被引:2,自引:0,他引:2
Andrew D. Richardson Ryan S. Anderson M. Altaf Arain Alan G. Barr Gil Bohrer Guangsheng Chen Jing M. Chen Philippe Ciais Kenneth J. Davis Ankur R. Desai Michael C. Dietze Danilo Dragoni Steven R. Garrity Christopher M. Gough Robert Grant David Y. Hollinger Hank A. Margolis Harry McCaughey Mirco Migliavacca Russell K. Monson J. William Munger Benjamin Poulter Brett M. Raczka Daniel M. Ricciuto Alok K. Sahoo Kevin Schaefer Hanqin Tian Rodrigo Vargas Hans Verbeeck Jingfeng Xiao Yongkang Xue 《Global Change Biology》2012,18(2):566-584
Phenology, by controlling the seasonal activity of vegetation on the land surface, plays a fundamental role in regulating photosynthesis and other ecosystem processes, as well as competitive interactions and feedbacks to the climate system. We conducted an analysis to evaluate the representation of phenology, and the associated seasonality of ecosystem‐scale CO2 exchange, in 14 models participating in the North American Carbon Program Site Synthesis. Model predictions were evaluated using long‐term measurements (emphasizing the period 2000–2006) from 10 forested sites within the AmeriFlux and Fluxnet‐Canada networks. In deciduous forests, almost all models consistently predicted that the growing season started earlier, and ended later, than was actually observed; biases of 2 weeks or more were typical. For these sites, most models were also unable to explain more than a small fraction of the observed interannual variability in phenological transition dates. Finally, for deciduous forests, misrepresentation of the seasonal cycle resulted in over‐prediction of gross ecosystem photosynthesis by +160 ± 145 g C m?2 yr?1 during the spring transition period and +75 ± 130 g C m?2 yr?1 during the autumn transition period (13% and 8% annual productivity, respectively) compensating for the tendency of most models to under‐predict the magnitude of peak summertime photosynthetic rates. Models did a better job of predicting the seasonality of CO2 exchange for evergreen forests. These results highlight the need for improved understanding of the environmental controls on vegetation phenology and incorporation of this knowledge into better phenological models. Existing models are unlikely to predict future responses of phenology to climate change accurately and therefore will misrepresent the seasonality and interannual variability of key biosphere–atmosphere feedbacks and interactions in coupled global climate models. 相似文献