Site-specific conformational determination in thermal unfolding studies of helical peptides using vibrational circular dichroism with isotopic substitution.
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Understanding the detailed mechanism of protein folding requires dynamic, site-specific stereochemical information. The short time response of vibrational spectroscopies allows evaluation of the distribution of populations in rapid equilibrium as the peptide unfolds. Spectral shifts associated with isotopic labels along with local stereochemical sensitivity of vibrational circular dichroism (VCD) allow determination of the segment sequence of unfolding. For a series of alanine-rich peptides that form alpha-helices in aqueous solution, we used isotopic labeling and VCD to demonstrate that the alpha-helix noncooperatively unwinds from the ends with increasing temperature. For these blocked peptides, the C-terminal is frayed at 5 degrees C. Ab initio level theoretical simulations of the IR and VCD band shapes are used to analyze the spectra and to confirm the conformation of the labeled components. The VCD signals associated with the labeled residues are amplified by coupling to the nonlabeled parts of the molecule. Thus small labeled segments are detectable and stereochemically defined in moderately large peptides in this report of site-specific peptide VCD conformational analysis. (+info)
Multicentric study on malignant pleural mesothelioma and non-occupational exposure to asbestos.
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Insufficient evidence exists on the risk of pleural mesothelioma from non-occupational exposure to asbestos. A population-based case-control study was carried out in six areas from Italy, Spain and Switzerland. Information was collected for 215 new histologically confirmed cases and 448 controls. A panel of industrial hygienists assessed asbestos exposure separately for occupational, domestic and environmental sources. Classification of domestic and environmental exposure was based on a complete residential history, presence and use of asbestos at home, asbestos industrial activities in the surrounding area, and their distance from the dwelling. In 53 cases and 232 controls without evidence of occupational exposure to asbestos, moderate or high probability of domestic exposure was associated with an increased risk adjusted by age and sex: odds ratio (OR) 4.81, 95% confidence interval (CI) 1.8-13.1. This corresponds to three situations: cleaning asbestos-contaminated clothes, handling asbestos material and presence of asbestos material susceptible to damage. The estimated OR for high probability of environmental exposure (living within 2000 m of asbestos mines, asbestos cement plants, asbestos textiles, shipyards, or brakes factories) was 11.5 (95% CI 3.5-38.2). Living between 2000 and 5000 m from asbestos industries or within 500 m of industries using asbestos could also be associated with an increased risk. A dose-response pattern appeared with intensity of both sources of exposure. It is suggested that low-dose exposure to asbestos at home or in the general environment carries a measurable risk of malignant pleural mesothelioma. (+info)
The Skn7 response regulator of Saccharomyces cerevisiae interacts with Hsf1 in vivo and is required for the induction of heat shock genes by oxidative stress.
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The Skn7 response regulator has previously been shown to play a role in the induction of stress-responsive genes in yeast, e.g., in the induction of the thioredoxin gene in response to hydrogen peroxide. The yeast Heat Shock Factor, Hsf1, is central to the induction of another set of stress-inducible genes, namely the heat shock genes. These two regulatory trans-activators, Hsf1 and Skn7, share certain structural homologies, particularly in their DNA-binding domains and the presence of adjacent regions of coiled-coil structure, which are known to mediate protein-protein interactions. Here, we provide evidence that Hsf1 and Skn7 interact in vitro and in vivo and we show that Skn7 can bind to the same regulatory sequences as Hsf1, namely heat shock elements. Furthermore, we demonstrate that a strain deleted for the SKN7 gene and containing a temperature-sensitive mutation in Hsf1 is hypersensitive to oxidative stress. Our data suggest that Skn7 and Hsf1 cooperate to achieve maximal induction of heat shock genes in response specifically to oxidative stress. We further show that, like Hsf1, Skn7 can interact with itself and is localized to the nucleus under normal growth conditions as well as during oxidative stress. (+info)
Investigation of the association between excess winter mortality and socio-economic deprivation.
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BACKGROUND: Excess winter mortality is higher in England and Wales than in other European countries with similar or lower average winter temperatures. It might be expected that excess winter mortality would be higher in areas with greater socio-economic deprivation, and if this were so preventive interventions could be directed at populations in these areas. The association between deprivation and excess winter mortality has not been adequately investigated in the past. The aim of this study was to look at the association between excess winter mortality and socio-economic deprivation, so that policy decisions to reduce this excess mortality could be appropriately directed. METHODS: Super Profile groups derived from the 1991 Census were used as a measure of socio-economic status. The age-standardized excess winter death index (EWDI) was calculated for each Super Profile group, for the population of Bradford. The EWDI was also calculated for the manufacturing districts (Office for National Statistics area classification), a relatively deprived group, and compared with that for England and Wales. RESULTS: No significant trend was found in age-standardized excess winter mortality across the Super Profile groups. The manufacturing districts had a similar EWDI to the national value. CONCLUSION: Excess winter mortality is not associated with deprivation. Further research to identify the important aetiological factors and appropriate interventions to reduce excess winter mortality is needed. (+info)
Effects of short episodes of heat stress on flower production and fruit-set of groundnut (Arachis hypogaea L.).
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Groundnuts (Arachis hypogaea L.) are an important crop of the semi-arid tropics where they are often exposed to maximum temperatures of > 40 degrees C for short periods during the growing season. The objectives of this study were to determine: (i) the effects of short periods of exposure to high temperature on flower production (FN), the proportion of flowers forming fruits (fruit-set) and the number of pegs and pods per plant (RNt); (ii) whether fruit-set is affected by high temperature during different periods of daylight in each diurnal cycle; and (iii) whether responses to temperature were qualitative or quantitative. Plants of cv. ICGV 86015 were grown in controlled environments at a day/night temperature of 28/22 degrees C from sowing until 9 d after flowering (DAF). Then, cohorts of plants were: (a) exposed to day temperature of 28, 34, 42 or 48 degrees C for 2, 4 or 6 d; or were (b) exposed to 34, 42 or 48 degrees C for 6 d either throughout a 12 h day (08.00 to 20.00 h, WD), or only during the first 6 h (AM) or second 6 h (PM) of the day. Values of RNt were significantly reduced by high temperature, by duration of exposure, and by timing of exposure. Variation in FN was quantitatively related to floral bud temperatures during the day over the range 28-43 degrees C. In contrast, only floral bud temperatures > 36 degrees C during AM and WD significantly reduced fruit-set and hence RNt, whereas high PM temperature had no effect on fruit-set. These findings indicate that the response of RNt to day temperature is quantitative and can be modelled by combining the responses of FN and fruit-set to temperature. (+info)
A two-dimensional protein gel electrophoresis study of the heat stress response of Bacillus subtilis cells during sporulation.
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The heat resistance of spores of Bacillus subtilis formed at 30 degrees C was enhanced by pretreatment at 48 degrees C for 30 min, 60 min into sporulation, for all four strains examined. High-resolution two-dimensional gel electrophoresis showed the generation and/or overexpression of 60 proteins, 11 of which were specific to heat shock, concurrent to this acquired thermotolerance. The greatest number of new proteins was observed between 30 and 60 min after heat shock, and the longer the time between exponential growth and heat treatment, the fewer differences were observed on corresponding protein profiles. The time at which heating produced the maximum increase in spore resistance and the most new proteins on two-dimensional gels occurred before alkaline phosphatase and dipicolinic acid production and corresponded to stage I or II of sporulation. The stress proteins formed disappeared later in sporulation, suggesting that heat shock proteins increase spore heat resistance by altering spore structure rather than by repairing heat damage during germination and outgrowth. (+info)
Developmental control of stress stimulons in Streptomyces coelicolor revealed by statistical analyses of global gene expression patterns.
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Stress-induced regulatory networks coordinated with a procaryotic developmental program were revealed by two-dimensional gel analyses of global gene expression. Four developmental stages were identified by their distinctive protein synthesis patterns using principal component analysis. Statistical analyses focused on five stress stimulons (induced by heat, cold, salt, ethanol, or antibiotic shock) and their synthesis during development. Unlike other bacteria, for which various stresses induce expression of similar sets of protein spots, in Streptomyces coelicolor heat, salt, and ethanol stimulons were composed of independent sets of proteins. This suggested independent control by different physiological stress signals and their corresponding regulatory systems. These stress proteins were also under developmental control. Cluster analysis of stress protein synthesis profiles identified 10 different developmental patterns or "synexpression groups." Proteins induced by cold, heat, or salt shock were enriched in three developmental synexpression groups. In addition, certain proteins belonging to the heat and salt shock stimulons were coregulated during development. Thus, stress regulatory systems controlling these stimulons were implicated as integral parts of the developmental program. This correlation suggested that thermal shock and salt shock stress response regulatory systems either allow the cell to adapt to stresses associated with development or directly control the developmental program. (+info)
Identification of enzymes homologous to isocitrate dehydrogenase that are involved in coenzyme B and leucine biosynthesis in methanoarchaea.
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Two putative Methanococcus jannaschii isocitrate dehydrogenase genes, MJ1596 and MJ0720, were cloned and overexpressed in Escherichia coli, and their gene products were tested for the ability to catalyze the NAD- and NADP-dependent oxidative decarboxylation of DL-threo-3-isopropylmalic acid, threo-isocitrate, erythro-isocitrate, and homologs of threo-isocitrate. Neither enzyme was found to use any of the isomers of isocitrate as a substrate. The protein product of the MJ1596 gene, designated AksF, catalyzed the NAD-dependent decarboxylation of intermediates in the biosynthesis of 7-mercaptoheptanoic acid, a moiety of methanoarchaeal coenzyme B (7-mercaptoheptanylthreonine phosphate). These intermediates included (-)-threo-isohomocitrate [(-)-threo-1-hydroxy-1,2, 4-butanetricarboxylic acid], (-)-threo-iso(homo)(2)citrate [(-)-threo-1-hydroxy-1,2,5-pentanetricarboxylic acid], and (-)-threo-iso(homo)(3)citrate [(-)-threo-1-hydroxy-1,2, 6-hexanetricarboxylic acid]. The protein product of MJ0720 was found to be alpha-isopropylmalate dehydrogenase (LeuB) and was found to catalyze the NAD-dependent decarboxylation of one isomer of DL-threo-isopropylmalate to 2-ketoisocaproate; thus, it is involved in the biosynthesis of leucine. The AksF enzyme proved to be thermostable, losing only 10% of its enzymatic activity after heating at 100 degrees C for 10 min, whereas the LeuB enzyme lost 50% of its enzymatic activity after heating at 80 degrees C for 10 min. (+info)