Redox reactivity of the heme Fe<Superscript>3+</Superscript>/Fe<Superscript>2+</Superscript> couple in native myoglobins and mutants with peroxidase-like activity

The reaction enthalpy and entropy for the one-electron reduction of the ferric heme in horse heart and sperm whale aquometmyoglobins (Mb) have been determined exploiting a spectroelectrochemical approach. Also investigated were the T67R, T67K, T67R/S92D and T67R/S92D Mb-H variants (the latter containing a protoheme-l-histidine methyl ester) of sperm whale Mb, which feature peroxidase-like activity. The reduction potential (E°′) in all species consists of an enthalpic term which disfavors Fe³⁺ reduction and a larger entropic contribution which instead selectively stabilizes the reduced form. This behavior differs from that of the heme redox enzymes and electron transport proteins investigated so far. The reduction thermodynamics in the series of sperm whale Mb variants show an almost perfect enthalpy–entropy compensation, indicating that the mutation-induced changes in are dominated by reduction-induced solvent reorganization effects. The modest changes in E°′ originate from the enthalpic effects of the electrostatic interactions of the heme with the engineered charged residues. The small influence that the mutations exert on the reduction potential of myoglobin suggests that the increased peroxidase activity of the variants is not related to changes in the redox reactivity of the heme iron, but are likely related to a more favored substrate orientation within the distal heme cavity.     

Topological research on diamagnetic susceptibilities of organic compounds

A novel molecular connectivity index, , based on the adjacency matrix of molecular graphs and novel atomic valence connectivities, , for predicting the molar diamagnetic susceptibilities of organic compounds is proposed. The is defined as: , where and E_i are the atomic valence connectivity and the valence orbital energy of atom i, respectively. A good QSPR model for molar diamagnetic susceptibilities can be constructed from and using multivariate linear regression (MLR). The correlation coefficient r, standard error, and average absolute deviation of the MLR model are 0.9918, 5.56 cgs, and 4.26 cgs, respectively, for the 721 organic compounds tested (training set). Cross-validation using the leave-one-out method demonstrates that the MLR model is highly reliable statistically. Using the MLR model, the average absolute deviations of the predicted values of molar diamagnetic susceptibility of another 360 organic compounds (test set) is 4.34 cgs. The results show that the current method is more effective than literature methods for estimating the molar diamagnetic susceptibility of an organic compound. The MLR method thus provides an acceptable model for the prediction of molar diamagnetic susceptibilities of organic compounds. Figure Plot of calculated vs experimental values of molar diamagnetic susceptibilities using the multivariate linear regression (MLR) model (Eq. 8)     

Effects of common soil anions and pH on the uptake and accumulation of perchlorate in lettuce

A mechanistic understanding of perchlorate () entry into plants is important for establishing the human health risk associated with consumption of contaminated produce and for assessing the effectiveness of phytoremediation. To determine whether common soil anions affect uptake and accumulation in higher plants, a series of competition experiments using lettuce (Lactuca sativa L.) were conducted between (50 nM) and (4–12 mM), (1–10 mM), or Cl⁻ (5–15 mM) in hydroponic solution. The effects of (0–5 mM) and pH (5.5–7.5) on uptake were also examined. Increasing in solution significantly reduced the amount of taken up by green leaf, butter head, and crisphead lettuces. Sulfate and Cl⁻ had no significant effects on uptake in lettuce over the concentrations tested. Increasing pH significantly reduced the amount of taken up by crisphead and green leaf lettuces, whereas increasing significantly reduced uptake in butter head lettuce. The inhibition by across all lettuce genotypes suggests that may share an ion carrier with , and the decrease in uptake with increasing pH or provides macroscopic evidence for cotransport across the plasma membrane.     

Kinetics and mechanistic studies of the reactions of metleghemoglobin, ferrylleghemoglobin, and nitrosylleghemoglobin with reactive nitrogen species

It is now established that nitrogen monoxide is produced not only in animals but also in plants. However, much less is known about the pathways of generation and the functions of in planta. One of the possible targets of is leghemoglobin (Lb), the hemoprotein found in high concentrations in the root nodules of legumes that establish a symbiosis with nitrogen-fixing bacteria. In analogy to hemoglobin and myoglobin, we have shown that different forms of Lb react not only with , but also with so-called reactive nitrogen species derived from it, among others peroxynitrite and nitrite. Because of the wider active-site pocket, the rate constants measured in this work for and for nitrite binding to metLb are 1 order of magnitude larger than the corresponding values for binding of these species to metmyoglobin and methemoglobin. Moreover, we showed that reactive nitrogen species are able to react with two forms of Lb that are produced in vivo but that cannot bind oxygen: ferrylLb is reduced by and nitrite, and nitrosylLb is oxidized by peroxynitrite. The second-order rate constants of these reactions are on the order of 10², 10⁶, and 10⁵ M⁻¹ s⁻¹, respectively. In all cases, the final reaction product is metLb, a further Lb form that has been detected in vivo. Since a specific reductase is active in nodules, which reduces metLb, reactive nitrogen species could contribute to the recycling of these inactive forms to regenerate deoxyLb, the oxygen-binding form of Lb.     

Inheritance of foliar stable carbon isotope discrimination and third-year height in Pinus taeda clones on contrasting sites in Florida and Georgia

Quantifying foliar stable carbon isotope discrimination (Δ) is a powerful approach for understanding genetic variation in gas exchange traits in large populations. The genetic architecture of Δ and third-year height is described for more than 1,000 clones of Pinus taeda tested on two contrasting sites. for Δ was 0.14 (±0.03), 0.20 (±0.07), and 0.09 (±0.04) at Florida, Georgia, and across sites, respectively. for stable carbon isotope discrimination ranged from 0.25 (±0.03) at the Florida site to 0.33 (±0.03) at the Georgia site, while the across-site estimate of was 0.19 (±0.02). For third-year height, ranged from 0.13 (±0.05) at the Georgia site to 0.20 (±0.06) at the Florida site with an across-site estimate of 0.09 (±0.05). Broad-sense heritability estimates for third-year height were 0.23 (±0.03), 0.28 (±0.03), and 0.13 (±0.02) at the Florida site, Georgia site, and across sites, respectively. Type B total genetic correlation for Δ was 0.70 ± 0.06, indicating that clonal rankings were relatively stable across sites, while for third-year height, rankings of clones were more unstable across the two trials . Third-year height and Δ were negatively correlated at the parental , full-sib family , and clonal levels, suggesting that genetic variation for Δ in P. taeda may be a result of differences in photosynthetic capacity. We conclude that Δ may be a useful selection trait to improve water-use efficiency and for guiding deployment decisions in P. taeda.     

Phase Transitions of Frozen Camu-camu (Myrciaria dubia (H.B.K.) McVaugh) Pulp: Effect of Cryostabilizer Addition

Camu-camu is a tropical fruit with very high vitamin C content and commercialized as frozen pulp. Enthalpies of freezing, temperatures of the onset of ice melting, and glass transition temperatures of the maximally freeze-concentrated phase () of camu-camu pulp and of samples containing maltodextrin (DE20) and sucrose were measured by differential scanning calorimetry. Maltodextrin exhibited the largest freeze stabilization potential, increasing from −58.2 °C (natural pulp) to −39.6 °C when 30% (w/w) maltodextrin DE 20 was added. Sucrose showed negligible effect on but enhanced considerably the freezing point depression and less amount of ice was formed.     

Effect of nickel on ROS content and antioxidative enzyme activities in wheat leaves

Influence of 100 μM Ni on growth, Ni accumulation,, H₂O₂ and lipid peroxides contents as well as the activities of superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), guaiacol peroxidase (POD) and glutathione peroxidase (GSH-Px) were studied in the leaves of wheat plants on the 3rd, 6th and 9th days after treatment. Exposure of the plants to Ni for only 3 days led to almost 200-fold increase in this metal concentration in the leaf tissue but later the rate of Ni accumulation was much slower. Length and fresh weight of the leaves were substantially reduced, up to 25% and 39%, respectively at the end of experiment. Visible symptoms of Ni toxicity: chlorosis and necrosis were observed following the 3rd day. Treatment with Ni resulted in the increase in and H₂O₂ contents in the leaves. Both showed their highest values, approximately 250% of those of the control, on the 3rd day and then their levels decreased but still markedly exceeded the control values. SOD and CAT activities decreased significantly in response to Ni treatment, however a several-fold increase in APX and POD activities was found. No significant changes in lipid peroxides content were observed in the leaves after Ni application. The activity of GSH-Px showed a 29% induction on the 3rd day. Our results indicated that despite prolonged increases in and H₂O₂ levels, oxidative damage, measured as the level of lipid peroxidation, did not occur in the leaves of Ni-treated wheat.     

Quantitative Γ-HCNCH: determination of the glycosidic torsion angle χ in RNA oligonucleotides from the analysis of CH dipolar cross-correlated relaxation by solution NMR spectroscopy

A novel NMR pulse sequence is introduced to determine the glycosidic torsion angle χ in ¹³C,¹⁵N-labeled oligonucleotides. The quantitative Γ-HCNCH measures the dipolar cross-correlated relaxation rates (pyrimidines) and (purines). Cross-correlated relaxation rates of a ¹³C,¹⁵N-labeled RNA 14mer containing a cUUCGg tetraloop were determined and yielded χ-angles that agreed remarkably well with data derived from the X-ray structure of the tetraloop. In addition, the method was applied to the larger stemloop D (SLD) subdomain of the Coxsackievirus B3 cloverleaf 30mer RNA and the effect of anisotropic rotational motion was examined for this molecule. It could be shown that the χ-angle determination especially for nucleotides in the anti conformation was very accurate and the method was ideally suited to distinguish between the syn and the anti-conformation of all four types of nucleotides. Electronic Supplementary Material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Engineering heme binding sites in monomeric rop

Heme ligands were introduced in the hydrophobic core of an engineered monomeric ColE1 repressor of primer (rop-S55) in two different layers of the heptad repeat. Mutants rop-L63M/F121H (layer 1) and rop-L56H/L113H (layer 3) were found to bind heme with a K _D of 1.1 ± 0.2 and 0.47 ± 0.07 μM, respectively. The unfolding of heme-bound and heme-free mutants, in the presence of guanidinium hydrochloride, was monitored by both circular dichroism and fluorescence spectroscopy. For the heme-bound rop mutants, the total free energy change was 0.5 kcal/mol higher in the layer 3 mutant compared with that in the layer1 mutant. Heme binding also stabilized these mutants by increasing the by 1.4 and 1.8 kcal/mol in rop-L63M/F121H and rop-L56H/L113H, respectively. The reduction potentials measured by spectroelectrochemical titrations were calculated to be −154 ± 2 mV for rop-56H/113H and −87.5 ± 1.2 mV for rop-L63M/F121H. The mutant designed to bind heme in a more buried environment (layer 3) showed tighter heme binding, a higher stability, and a different reduction potential compared with the mutant designed to bind heme in layer 1.     

A bias correction for estimates of effective population size based on linkage disequilibrium at unlinked gene loci*

Analysis of linkage disequilibrium (=mean squared correlation of allele frequencies at different gene loci) provides a means of estimating effective population size (N _e) from a single sample, but this method has seen much less use than the temporal method (which requires at least two samples). It is shown that for realistic numbers of loci and alleles, the linkage disequilibrium method can provide precision comparable to that of the temporal method. However, computer simulations show that estimates of N _e based on for unlinked, diallelic gene loci are sharply biased downwards ( in some cases) if sample size (S) is less than true N _e. The bias is shown to arise from inaccuracies in published formula for when S and/or N _e are small. Empirically derived modifications to for two mating systems (random mating and lifetime monogamy) effectively eliminate the bias (residual bias in % in most cases). The modified method also performs well in estimating N _e in non-ideal populations with skewed sex ratio or non-random variance in reproductive success. Recent population declines are not likely to seriously affect , but if N has recently increased from a bottleneck can be biased downwards for a few generations. These results should facilitate application of the disequilibrium method for estimating contemporary N _e in natural populations. However, a comprehensive assessment of performance of with highly polymorphic markers such as microsatellites is needed.The US Governmentȁ9s right to retain a non-exclusive, royalty-free license in and to any copyright is acknowledged.     

A Combined Patch-Clamp and Electrorotation Study of the Voltage- and Frequency-Dependent Membrane Capacitance Caused by Structurally Dissimilar Lipophilic Anions

Interactions of structurally dissimilar anionic compounds with the plasma membrane of HEK293 cells were analyzed by patch clamp and electrorotation. The combined approach provides complementary information on the lipophilicity, preferential affinity of the anions to the inner/outer membrane leaflet, adsorption depth and transmembrane mobility. The anionic species studied here included the well-known lipophilic anions dipicrylamine (DPA⁻), tetraphenylborate (TPB⁻) and [W₂(CO)₁₀(S₂CH)]⁻, the putative lipophilic anion and three new heterocyclic W(CO)₅ derivatives. All tested anions partitioned strongly into the cell membrane, as indicated by the capacitance increase in patch-clamped cells. The capacitance increment exhibited a bell-shaped dependence on membrane voltage. The midpoint potentials of the maximum capacitance increment were negative, indicating the exclusion of lipophilic anions from the outer membrane leaflet. The adsorption depth of the large organic anions DPA⁻, TPB⁻ and increased and that of W(CO)₅ derivatives decreased with increasing concentration of mobile charges. In agreement with the patch-clamp data, electrorotation of cells treated with DPA⁻ and W(CO)₅ derivatives revealed a large dispersion of membrane capacitance in the kilohertz to megahertz range due to the translocation of mobile charges. In contrast, in the presence of TPB⁻ and no mobile charges could be detected by electrorotation, despite their strong membrane adsorption. Our data suggest that the presence of oxygen atoms in the outer molecular shell is an important factor for the fast translocation ability of lipophilic anions.     

Sources of variability in spotted owl population growth rate: testing predictions using long-term mark–recapture data

For long-lived iteroparous vertebrates that annually produce few young, life history theory predicts that reproductive output (R) and juvenile survival should influence temporal variation in population growth rate (λ) more than adult survival does. We examined this general prediction using 15 years of mark–recapture data from a population of California spotted owls (Strix occidentalis occidentalis). We found that survival of individuals ≥1 year old (ϕ) exhibited much less temporal variability , where CV is coefficient of variation, than R and that R was strongly influenced by environmental stochasticity. Although λ was most sensitive ( ; log-transformed sensitivity) to ϕ and much less sensitive to either R or juvenile survival (survival rate of owls from fledging to 1 year old; ), we estimated that R contributed as much as ϕ to the observed annual variability in λ. The contribution of juvenile survival to variability in λ was proportional to its These results are consistent with the hypothesis that natural selection may have favored the evolution of longevity in spotted owls as a strategy to increase the probability of experiencing favorable years for reproduction. Our finding that annual weather patterns that most affected R (temperature and precipitation during incubation) and ϕ (conditions during winter related to the Southern Oscillation Index) were equally good at explaining temporal variability in λ supports the conclusion that R and ϕ were equally responsible for variability in λ. Although currently accepted conservation measures for spotted owl populations attempt to enhance survival, our results indicated that conservation measures that target R may be as successful, as long as actions do not reduce ϕ.     

Use of isotopic signatures to assess the food web in a tropical shallow marine ecosystem of Southeastern Brazil

A dual isotope approach was used to assess the relative importance of terrestrial vegetation detritus and other primary producers in the trophic web of Flamengo Sound (Ubatuba, SP), SE Brazil, surrounded by the Atlantic Rain Forest. Primary producers showed distinct C signatures and the observed values suggest that little terrestrial or bulk sediment organic matter enter the food web of the sound. Suspended particulate organic matter (POM, supports the bulk of the consumers, with some contribution by macroalgae . Consumers C values ranged from −17.4 to . At least three trophic levels were detectable in the food web. The N value of POM was , while that of sediment and detritus was . The N values of suspension feeding benthic invertebrates were 8.2–, deposit feeders 8.3–, and carnivores 10.7–. Values for fishes were for detritivore, 11.4– for benthic feeders, 12.4– for zooplanktivores, and for piscivores/benthic invertebrate feeders. Squid mean value was . There is a reasonable agreement between feeding habits information from the literature and N values from this study. In the sound, the first and second trophic steps seem to be about 1– higher than those of similar organisms studied in temperate waters and this may reflect an input of allochtonous anthropogenic nitrogen enriched in ¹⁵N from human activities.     

Factors affecting denitrification in agricultural headwater streams in Northeast Ohio,USA

As a result of increased anthropogenic nitrogen (N) loading in surface waters of agricultural watersheds, there is enhanced interest to understand and quantify N removal mechanisms. Denitrification, an important N removal mechanism in aquatic systems, may contribute to reducing N pollution in agricultural headwater streams. However, the key factors controlling this process in lotic systems remain unclear. The objective of our study was to examine the factors regulating rates of denitrification in the sediments of agricultural headwater streams in the mid-western USA. Denitrification rates were variable among streams and treatments (<0.1–28.0 μg N g AFDM⁻¹ h⁻¹) and on average, were higher than those reported for similar headwater streams. Carbon quantity and quality, and pH had no effect on denitrification, while temperature and nitrate ( ) concentrations had a positive effect on rates of denitrification. Specifically, controlled denitrification following Michaelis-Menten kinetics. We calculated a value of k_m (1.0 mg -N L^-1) that was comparable to other studies in aquatic sediments but was well below the median in-stream concentrations (5.2–17.4 mg -N L⁻¹) observed at the study sites. Despite high rates of denitrification, this removal mechanism is most likely saturated in the agricultural headwater streams we examined, suggesting that these systems are not effective at removing in-stream N. Handling editor: D. Ryder     

RBE of quasi-monoenergetic 60 MeV neutron radiation for induction of dicentric chromosomes in human lymphocytes

The production of dicentric chromosomes in human lymphocytes by high-energy neutron radiation was studied using a quasi-monoenergetic 60 MeV neutron beam. The average yield coefficient of the linear dose–response relationship for dicentric chromosomes was measured to be (0.146±0.016) Gy⁻¹. This confirms our earlier observations that above 400 keV, the yield of dicentric chromosomes decreases with increasing neutron energy. Using the linear-quadratic dose–response relationship for dicentric chromosomes established in blood of the same donor for ⁶⁰Co γ-rays as a reference radiation, an average maximum low-dose RBE (RBE_M) of 14±4 for 60 MeV quasi-monoenergetic neutrons with a dose-weighted average energy of 41.0 MeV is obtained. A correction procedure was applied, to account for the low-energy continuum of the quasi-monoenergetic spectral neutron distribution, and the yield coefficient α for 60 MeV neutrons was determined from the measured average yield coefficient . For α, a value of (0.115±0.026) Gy⁻¹ was obtained corresponding to an RBE_M of 11±4. The present experiments extend earlier investigations with monoenergetic neutrons to higher energies.     

Genetic differentiation of Rosa canina (L.) at regional and continental scales

We investigated the genetic differentiation of Rosa canina at regional and continental scales to delineate provenance regions for seed collection. Using RAPD technique, we examined eight populations in Brandenburg (Germany) and 13 samples from Bavaria and Hungary. AMOVA and $\upphiWe investigated the genetic differentiation of Rosa canina at regional and continental scales to delineate provenance regions for seed collection. Using RAPD technique, we examined eight populations in Brandenburg (Germany) and 13 samples from Bavaria and Hungary. AMOVA and -statistics revealed a significant differentiation ( = 0.23, P < 0.0001) between accessions from Brandenburg and the two other regions. The interpopulation differentiation in Brandenburg was low ( = 0.13). We found the greatest interpopulation distance between Hungary and Brandenburg (Hoher Fl?ming, = 0.47) and the lowest between Dahmeland and Hoher Fl?ming (both Brandenburg, = 0.01). UPGMA dendrogram and PCA showed a clear disjunction between Brandenburg and other regions. Due to the outcrossing mating and seed dispersal system, the genetic variation within R. canina populations is moderately high (87% for Brandenburg, 76% for Bavaria and Hungary). The significant genetic differentiation at the continental scale suggests lower gene flow with increasing geographical distance and underlines the usefulness of regional provenances for planting.     

New complete orthonormal sets of hyperspherical harmonics and their one-range addition and expansion theorems

In this study, the complete orthonormal sets of - momentum space orbitals (where α=1,0,−1, −2,...) obtained from the -ETO in coordinate representation (I.I. Guseinov, J. Mol. Model., 9 (2003) 135) are reduced to the complete orthonormal sets of hyperspherical harmonics (HSH) by means of a Fock transformation of the radial momentum to an angular variable. It is shown that the group of transformations is the four-dimensional rotation group O(4) and, therefore, the HSH presented in this work are the complete orthonormal sets of functions. For these functions, the one-range addition and expansion theorems are obtained. The formulae for HSH and their addition and expansion theorems derived in this work can be used to evaluate the multicenter integrals that arise when exponential-type basis functions are used in atomic and molecular calculations.     

Theoretical study of AlH(2+)n (n=1-7) dications

The structures and stability of 1–7 dications were calculated at the ab initio MP2/aug-cc-pVTZ level of theory. The dications AlH²⁺ 1 and 2 were characterized to be unstable thermodynamically. However, these and the stable dications, 3–7 have considerable kinetic barriers for deprotonation. Each of the structures 3–7 contains one or more two-electron three-center (2e–3c) bonds. Aluminum atoms of these dications carry most of the positive charges, as indicated by NBO charge calculations.Dedicated to Professor Dr. Paul von Ragué Schleyer on the occasion of his 75th birthday