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991.
Mulkidjanian Armen Y. Hochkoeppler Alejandro Zannoni Davide Drachev Lel A. Melandri Bruno Andrea Venturoli Giovanni 《Photosynthesis research》1998,56(1):75-82
Light-dependent reduction of cystine disulfide bonds results in activation of several of the enzymes of photosynthetic carbon metabolism within the chloroplast. Tertiary structure modeling suggests that the redox-sensitivity of the chloroplast malate dehydrogenase (EC 1.1.1.82) is due to disulfide crosslinking of the carbon substrate and nucleotide-binding domains. Consistent with this suggestion, introduction of Cys residues in opposition to one another on the two domains of the Escherichia coli enzyme results in redox-sensitivity [Muslin EH et al. (1995) Biophys J 68: 2218-2223]. We have now substituted Cys residues into the bacterial malate dehydrogenase (EC 1.1.1.37) in positions that correspond more exactly to those postulated to be responsible for the redox-sensitivity of the chloroplast enzyme. The introduction of one pair of Cys residues renders the enzyme redox-sensitive, but the introduction of the alternate pair does not. Energy minimization calculations suggest that the difference in redox-sensitivity is consistent with differences in the energy required for formation of the disulfide bond. 相似文献
992.
993.
J. Andy Rakestraw Stephen L. Sazinsky Andrea Piatesi Eugene Antipov K. Dane Wittrup 《Biotechnology and bioengineering》2009,103(6):1192-1201
Because of its eukaryotic nature, simple fermentation requirements, and pliable genetics, there have been many attempts at improving recombinant protein production in Saccharomyces cerevisiae. These strategies typically involve altering the expression of a native protein thought to be involved in heterologous protein trafficking. Usually, these approaches yield three‐ to tenfold improvements over wild‐type strains and are almost always specific to one type of protein. In this study, a library of mutant alpha mating factor 1 leader peptides (MFα1pp) is screened for the enhanced secretion of a single‐chain antibody. One of the isolated mutants is shown to enhance the secretion of the scFv up to 16‐fold over wild type. These leaders also confer a secretory improvement to two other scFvs as well as two additional, structurally unrelated proteins. Moreover, the improved leader sequences, combined with strain engineering, allow for a 180‐fold improvement over previous reports in the secretion of full‐length, functional, glycosylated human IgG1. The production of full‐length IgG1 at milligram per liter titers in a simple, laboratory‐scale system will significantly expedite drug discovery and reagent synthesis while reducing antibody cloning, production, and characterization costs. Biotechnol. Bioeng. 2009;103: 1192–1201. © 2009 Wiley Periodicals, Inc. 相似文献
994.
Diana Poli Andrea Caglieri Matteo Goldoni Anna F. Castoldi Teresa Coccini Elisa Roda Annabella Vitalone Sandra Ceccatelli Antonio Mutti 《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》2009,877(8-9):773-783
A simple and reliable solid phase microextraction/gas chromatography–mass spectrometry (SPME/GC–MS) method was developed for the single-step determination of PCBs 126 and 153 in rat brain and serum, using liquid/liquid and solid phase extraction (SPE) as reference techniques. The multi-factor categorical experimental design used to study simultaneously the main parameters and their interactions affecting the efficiency of the method, showed that the use of an 85 μm PA exposed at 100 °C for 40 min was the optimum sampling condition for both PCBs. SPME was then validated by studying its linear dynamic (over two orders of magnitude), limits of detection (brain: 2 ng/g, serum: 0.2 ng/g) and analytical precision that was within 9% for SPME in both brain and serum. Finally, the method was used to determine the brain and blood target dose in mothers and pups after oral exposure of the mothers. 相似文献
995.
David Giraldi Mattia de’Michieli Vitturi Mattia Zaramella Andrea Marion Renato Iannelli 《Ecological Engineering》2009,35(2):265-273
Typical unsteady unsaturated conditions can profoundly affect the hydrodynamics of vertical subsurface flow constructed wetlands. In this study we analyzed the hydrodynamics of a 33 m2 vertical flow pilot plant, treating municipal secondary effluents. Three different saturation conditions were analyzed under several constant flux regimes: complete saturation, partial saturation with the free water table 20 cm over the bottom of the bed, and complete drainage. Tracer tests were performed in steady state conditions by dosing rhodamine WT as square input signals. Breakthrough curves were analyzed by means of both a classical residence time distribution analysis and an originally developed numerical plug-flow model with longitudinal dispersion adapted to the unsaturated conditions. We found that the degree of global mixing in the vertical flow constructed wetland increased as the water content increased; this effect was controlled by the hydraulic residence time of the system. Conversely, the degree of local mixing was inversely affected by water content; the dispersivity was 4.5, 10, and 14 cm for fully saturated, partially saturated and draining conditions, respectively. We explain the dependency of dispersivity on water content in physical terms; however, further studies are needed to mathematically include this relationship in numerical models that describe the behaviour of vertical flow constructed wetlands. 相似文献
996.
Sara Alberghini Elisa Polone Viviana Corich Milena Carlot Flavio Seno Antonio Trovato & Andrea Squartini 《FEMS microbiology letters》2009,292(2):149-161
Cell-to-cell bacterial communication via diffusible signals is addressed and the conceptual framework in which quorum sensing is usually described is evaluated. By applying equations ruling the physical diffusion of the autoinducer molecules, one can calculate the gradient profiles that would occur either around a single cell or at the center of volumes of increasing size and increasing cell densities. Water-based matrices at 25 °C and viscous biofilms at colder temperatures are compared. Some basic consequences relevant for the field of microbial signalling arise. As regards induction, gradient-mixing dynamics between as little as two cells lying at a short distance appears to be sufficient for the buildup of a concentration reaching the known thresholds for quorum sensing. A straight line in which the highest concentrations occur is also created as a consequence of the gradient overlap geometry, providing an additional signal information potentially useful for chemotactic responses. In terms of whole population signalling, it is shown how the concentration perceived by a cell in the center is critically dependent not only on the cell density but also on the size of the biofilm itself. Tables and formulas for the practical prediction of N -acyl homoserine lactones concentrations at desired distances in different cell density biofilms are provided. 相似文献
997.
To better understand the mechanism(s) of leukocyte Ca(2+) signaling, we have studied the intracellular locations of two Ca(2+)-mobilizing receptors, the inositol 1,4,5-trisphosphate receptor and ryanodine receptor, by immunofluorescence microscopy. Our results show that localization differs not only between receptor classes within a cell, but among leukocyte types as well. We also illustrate the importance of preserving labile cellular filaments in maintaining cell integrity by fixation with the Safiejko-Mroczka and Bell protocol, because conventional fixation methods distort receptor patterns. We suggest that the observed differences influence intracellular Ca(2+) signaling. 相似文献
998.
Rafael Rodrigues Silva Rafael M. Mariante Andrea Alice Silva Ana Luiza Barbosa dos Santos Ester Roffê Helton Santiago Ricardo Tostes Gazzinelli Joseli Lannes-Vieira 《PloS one》2015,10(2)
The inflammatory cytokine interferon-gamma (IFNγ) is crucial for immunity against intracellular pathogens such as the protozoan parasite Trypanosoma cruzi, the causative agent of Chagas disease (CD). IFNγ is a pleiotropic cytokine which regulates activation of immune and non-immune cells; however, the effect of IFNγ in the central nervous system (CNS) and astrocytes during CD is unknown. Here we show that parasite persists in the CNS of C3H/He mice chronically infected with the Colombian T. cruzi strain despite the increased expression of IFNγ mRNA. Furthermore, most of the T. cruzi-bearing cells were astrocytes located near IFNγ+ cells. Surprisingly, in vitro experiments revealed that pretreatment with IFNγ promoted the infection of astrocytes by T. cruzi increasing uptake and proliferation of intracellular forms, despite inducing increased production of nitric oxide (NO). Importantly, the effect of IFNγ on T. cruzi uptake and growth is completely blocked by the anti-tumor necrosis factor (TNF) antibody Infliximab and partially blocked by the inhibitor of nitric oxide synthesis L-NAME. These data support that IFNγ fuels astrocyte infection by T. cruzi and critically implicate IFNγ-stimulated T. cruzi-infected astrocytes as sources of TNF and NO, which may contribute to parasite persistence and CNS pathology in CD. 相似文献
999.
Antonio Pacilli Olga Lamacchia Andrea Fontana Massimiliano Copetti Mauro Cignarelli Vincenzo Trischitta Salvatore De Cosmo 《PloS one》2015,10(4)
Background
To investigate prospectively the relationship between target values of glycated hemoglobin, blood pressure and LDL-cholesterol, as considered in a combined fashion, and all-cause mortality in patients with type 2 diabetes mellitus.Methods
Two cohorts of patients with type 2 diabetes mellitus, the Gargano Mortality Study (n=810) and the Foggia Mortality Study (n=929), were investigated. A weighted target risk score was built as a weight linear combination of the recommended targets reached by each patient.Results
In the Gargano Mortality Study and in the Foggia Mortality Study (mean follow up=7.4 and 5.5 years, respectively), 161 (19.9%) and 220 (23.7%) patients died, with an age and sex adjusted annual incidence rate of 2.1 and 2.8 per 100 person-years, respectively. In both study samples the weighted target risk score tended to be linearly associated with all-cause mortality (HR for one point increment=1.30, 95% CI: 1.11-1.53, p=0.001, and HR=1.08, 95% CI: 0.95-1.24, p=0.243, respectively). When the two cohorts were pooled and analyzed together, a clear association between weighted target risk score and all-cause mortality was observed (HR for one point increment=1.17, 95% CI:1.05-1.30, p=0.004). This counterintuitive association was no longer observable in a model including age, sex, body mass index, smoking habit, estimated glomerular filtration rate, albuminuria and anti-diabetic, anti-hypertensive and anti-dyslipidemic treatment as covariates (HR for one point increment=0.99, 95% CI: 0.87-1.12, p=0.852).Conclusions
In a real life clinical set of patients with type 2 diabetes mellitus, the combination of recommended target values of established cardiovascular risk factors is not associated with all-cause mortality. 相似文献1000.
Ylenia Ingrasciotta Janet Sultana Francesco Giorgianni Andrea Fontana Antonio Santangelo Daniele Ugo Tari Domenico Santoro Vincenzo Arcoraci Margherita Perrotta Luisa Ibanez Gianluca Trifirò 《PloS one》2015,10(4)