Mössbauer, EPR, and MCD studies of the C9S and C42S variants of Clostridium pasteurianum rubredoxin and MCD studies of the wild-type protein

Rubredoxins contain a mononuclear iron tetrahedrally coordinated by four cysteinyl sulfurs. We have studied the wild-type protein from Clostridium pasteurianum and two mutated forms, C9S and C42S, in the oxidized and reduced states, with Mössbauer, integer-spin EPR, and magnetic circular dichroism (MCD) spectroscopies. The Mössbauer spectra of the ferric C42S and C9S mutant forms yielded zero-field splittings, D=1.2?cm^?1, that are about 40% smaller than the D-value of the wild-type protein. The ⁵⁷Fe hyperfine coupling constants were found to be ca. 8% larger than those of the wild-type proteins. The present study also revealed that the ferric wild-type protein has δ=0.24±0.01?mm/s at 4.2?K rather than δ=0.32?mm/s as reported in the literature. The Mössbauer spectra of both dithionite-reduced mutant proteins revealed the presence of two ferrous forms, A and B. These forms have isomer shifts δ=0.79?mm/s at 4.2?K, consistent with tetrahedral Fe²⁺(Cys)₃(O-R) coordination. The zero-field splittings of the two forms differ substantially; we found D=?7±1?cm^?1, E/D=0.09 for form A and D=+6.2±1.3?cm^?1, E/D=0.15 for form B. Form A exhibits a well-defined integer-spin EPR signal; from studies at X- and Q-band we obtained g _z =2.08±0.01, which is the first measured g-value for any ferrous rubredoxin. It is known from X-ray crystallographic studies that ferric C42S rubredoxin is coordinated by a serine oxygen. We achieved 75% reduction of C42S rubredoxin by irradiating an oxidized sample at 77?K with synchrotron X-rays; the radiolytic reduction produced exclusively form A, suggesting that this form represents a serine-bound Fe²⁺ site. Studies in different buffers in the pH?6–9 range showed that the A:B ratios, but not the spectral parameters of A and B, are buffer dependent, but no systematic variation of the ratio of the two forms with pH was observed. The presence of glycerol (30–50% v/v) was found to favor the B form. Previous absorption and circular dichroism studies of reduced wild-type rubredoxin have suggested d-d bands at 7400, 6000, and 3700?cm^?1. Our low-temperature MCD measurements place the two high-energy transitions at ca. 5900 and 6300?cm^?1; a third d-d transition, if present, must occur with energy lower than 3300?cm^?1. The mutant proteins have d-d transitions at slightly lower energy, namely 5730, 6100?cm^?1 in form A and 5350, 6380?cm^?1 in form B.     

Sleep-Wake Cycle of Adolescents in Côte d'Ivoire: Influence of Age,Gender, Religion and Occupation

The human sleep-wake cycle is characterized by significant individual differences. Those differences in the sleep-wake cycle are partially heritable but are also influenced by environmental factors like the light/dark cycle or social habits. In this study we analyse for the first time the sleep-wake rhythm of adolescent pupils and working adolescents in a less industrialised country in West Africa near the equator. The aim of this study was to explore the sleep wake cycle in this geographical region, using Côte d'Ivoire as an example. Data collection took place between 2nd of March and 10th of June 2009. 588 adolescents (338 girls, 250 boys) between 10 and 15 years (mean?±?SD: 12.72?±?1.63) participated in this study. We collected data on the religion of the participants (Christian (N?=?159), Muslim (N?=?352), other/no religion (N?=?77)) and their occupation. Participants were either pupils attending school (N?=?336) or adolescents that were already working (N?=?252) and not attending school. The interviewer filled in the questionnaire. We found significant effects of age (p?<?0.001), gender (p?<?0.001), occupation (p?=?0.002), religion (p?<?0.001) and region (p?<?0.001). The midpoint of sleep was on average 1:26 (SD: 00:30) on weekdays and 1:37 (SD: 00:42) on weekend days. There are significant differences between weekdays and weekend days, but these were only small. Sleep duration suggests that adolescents in Côte d'Ivoire may gain sufficient sleep during week and weekend days, and thus, may live more in accordance with their own biological clock than adolescents in the northern hemisphere. In contrast, the data can be interpreted that adolescents live in a permanent ‘jetlag’. Factors may be the more continuous light/dark cycle in the tropics, low amount of ambient light and less electricity.     

Aphids (Hemiptera: Aphidoidea) of ornamental plants from São Carlos,São Paulo state,Brazil

A total of 25 aphid species were collected from 49 ornamental plant species in S?o Carlos-S?o Paulo, Brazil; 12 aphids were monophagous, four oligophagous and nine polyphagous. A total of 58 aphid-plant associations are recorded, 43 unknown from Brazil. Eucarazzia elegans (Ferrari, 1872) (Aphididae: Aphidinae: Macrosiphini) is recorded for the first time from Brazil, and Nectandra megapotamica Spreng. (Lauraceae) is recorded for the first time as host plant for Lizerius tuberculatus (E.E. Blanchard, 1939) (Drepanosiphidae: Drepanosiphinae: Lizeriini). We also describe the injuries caused by aphids to the ornamental plants.     

Darwin,Malthus, Süssmilch,and Euler: The Ultimate Origin of the Motivation for the Theory of Natural Selection

It is fairly well known that Darwin was inspired to formulate his theory of natural selection by reading Thomas Malthus’s Essay on the Principle of Population. In fact, by reading Darwin’s notebooks, we can even locate one particular sentence which started Darwin thinking about population and selection. What has not been done before is to explain exactly where this sentence – essentially Malthus’s ideas about geometric population growth – came from. In this essay we show that eighteenth century mathematician Leonhard Euler is responsible for this sentence, and in fact forms the beginning of the logical chain which leads to the creation of the theory of natural selection. We shall examine the fascinating path taken by a mathematical calculation, the many different lenses through which it was viewed, and the path through which it eventually influenced Darwin.     

Population structure of São Miguel Island, Azores: a surname study

The knowledge of a population structure may constitute a powerful tool for mapping genes underlying susceptibility to Mendelian and complex diseases. To obtain a better understanding of the population structure of S?o Miguel Island (Azorean Archipelago, Portugal), we carried out a surname survey using the surnames listed in the most recent telephone book (2001). We identified 1315 different surnames in a total of 27,621 subscribers. The frequency of the different surnames was used to calculate the following parameters: isonymy (I), random component of inbreeding (FST), genetic diversity according to Fisher (alpha), migration rate according to Karlin-McGregor (v) and Nei's genetic distance. Eleven localities were selected, according to population size and geographic distribution, for analysis using the above parameters. Our results show that 51% of Salga's population and 52% of Sete Cidades's population are represented by six and eight surnames, respectively. These figures demonstrate the effective isolation of these two small places, which are located at opposite ends of S?o Miguel Island. Salga, Achada, and Sete Cidades present the lowest values of Fisher's alpha, indicating less genetic diversity. In contrast, the capital, Ponta Delgada, presents the highest value of alpha (78.13), indicating more genetic diversity. Our data indicate that the clustering of the localities corresponds to the geographic features of the island, where localities close together tend to share similar surnames. In conclusion, the population of S?o Miguel is relatively homogeneous and may constitute an ideal model for genetic mapping studies.     

Overexpression of myoglobin and assignment of its amide,Cα and Cβ resonances

Summary Sperm whale apomyoglobin was expressed to high levels on minimal media and isotopically labeled with ¹³C and ¹⁵N nuclei. The isotopically labeled apoprotein was purified to homogeneity in a single step by reversed-phase chromatography and reconstituted with hemin and carbon monoxide gas for NMR analysis. Sequence-specific backbone ¹H^N, ¹⁵N and ¹³C as well as side-chain ¹³C resonance assignments have been made for over 90% of the amino acids in the carbon monoxide complex of the protein. Resonance assignments were made by analysis of a series of 3D triple resonance spectra measured on the uniformly labeled sample. These assignments will provide the basis for analyzing the effects of point site mutations on the structure, stability and dynamics of the protein in solution.     

Spread of Phlebotominae in temperate climates: province of Córdoba, Argentina

We report the presence of the competent vector for Leishmaniaspp, Migonemyia migonei, and theEvandromyia cortelezzii-sallesi complex south of its known distribution in the central temperate region of Argentina, in the province of Córdoba. The persistence of this phlebotomine in the northern border of the province, its association with a case of cutaneous leishmaniasis, and the new record in the outskirts of the city of Córdoba, the second most populated in the country, strengthens the need for regular vector surveillance and a case detection-sensitive health system in vulnerable regions, even in temperate climates.     

Sleep Patterns of Day-Working,Evening High-Schooled Adolescents of São Paulo,Brazil

Children who grow up in developing countries of the world must work to help financially support their families, and they must also attend school. We investigated the impact of work on the sleep of working vs. nonworking high school students. Twenty-seven São Paulo, Brazil, public high school students (eight male and eight female working students plus six nonworking female and five nonworking male students) 14–18 yrs of age who attended school Monday–Friday between 19:00 to 22:30 h participated. A comprehensive questionnaire about work and living conditions, health status, and diseases and their symptoms was also answered. The activity level and rest pattern (sleep at night and napping during the day) were continuously assessed by wrist actigraphy (Ambulatory Monitoring, USA). The main variables were analyzed by a two-factor ANOVA with application of the Tukey HSD test for multiple comparisons, and the length of sleep during weekdays vs. weekends was compared by Student t-test. Working students went to sleep earlier weekends [F_(1,23) = 6.1; p = 0.02] and woke up earlier work days than nonworking students [F_(1,23) = 17.3; p = 0.001]. The length of nighttime sleep during weekdays was shorter among all the working [F_(1,23) = 16.7; p < 0.001] than all the nonworking students. The sleep duration of boys was shorter than of girls during weekends [F_(1,23) = 10.8; p < 0.001]. During weekdays, the duration of napping by working and nonworking male students was shorter than nonworking female students. During weekdays, working girls took the shortest naps [F_(1,23) = 5.6; p = 0.03]. The most commonly reported sleep complaint during weekdays was difficulty waking up in the morning [F_(1,23) = 6.5; p = 0.02]. During weekdays, the self-perceived sleep quality of working students was worse than nonworking students [F_(1,23) = 6.2; p = 0.02]. The findings of this study show that work has negative effects on the sleep of adolescents, with the possible build-up of a chronic sleep debt with potential consequent impact on quality of life and school learning.     

Molecular properties of phosphoenolpyruvate carboxylase from C3, C3−C4 intermediate,and C4 Flaveria species

Phosphoenolpyruvate carboxylase (PEPCase; EC 4.1.1.31) from Flaveria trinervia Mohr (C₄), F. floridana Johnston (C₃–C₄), and F. cronquistii Powell (C₃) leaves were compared by electrotransfer blotting/enzyme-linked immunoassay (Western-blot analysis), mobility of the native enzyme in polyacrylamide gels and in isoelectric focusing (IEF) gels, peptide mapping, and in-vitro translation of RNA isolated from each plant. The PEPCases from the C₃ and C₃–C₄ plants were very similar to each other in terms of electrophoretic mobilities on gels and isoenzyme patterns on IEF gels, and identical in peptide mapping. Quantitative differences were noted, however, in that the C₃–C₄ intermediate plant contained more PEPCase overall and that the relative activity of individual isoenzymes shifted between the C₃ and C₃–C₄ intermediate PEPCases. The PEPCase from the C₄ plant had a different isoenzyme pattern, a different peptide map, and was far more abundant than the other two enzymes. Western blot analysis demonstrated the cross-reactivity of PEPCases from all three Flaveria species with antibody raised against maize PEPCase. The results provide evidence, at the molecular level, that supports the view of C₃–C₄ intermediate species as C₃-like plants with some C₄-like photosynthetic characteristics, but there are differences from the C₃ plant in the quantity and properties of the PEPCase from the C₃–C₄ intermediate plant.Abbreviations IEF isoelectric focusing - kDa kilodalton - PEPCase phosphoenolpyruvate carboxylase - Rubisco Ribulose-1,5-bisphosphate carboxylase/oxygenase