Heterometallic [AgFe3S4] ferredoxin variants: synthesis, characterization, and the first crystal structure of an engineered heterometallic iron–sulfur protein

Heterometallic [AgFe₃S₄] iron–sulfur clusters assembled in wild-type Pyrococcus furiosus ferredoxin and two variants, D14C and D14H, are characterized. The crystal structure of the [AgFe₃S₄] D14C variant shows that the silver(I) ion is indeed part of the cluster and is coordinated to the thiolate group of residue 14. Cyclic voltammetry shows one redox pair with a reduction potential of +220 mV versus the standard hydrogen electrode which is assigned to the [AgFe₃S₄]^2+/+ couple. The oxidized form of the [AgFe₃S₄] D14C variant is stable in the presence of dioxygen, whereas the oxidized forms of the [AgFe₃S₄] wild type and D14H variants convert to the [Fe₃S₄] ferredoxin form. The monovalent d ¹⁰ silver(I) ion stabilizes the [Fe₃S₄]^+/0 cluster fragment, as opposed to divalent d ¹⁰ metal ions, resulting in more than 0.4 V difference in reduction potentials between the silver(I) and, e.g., zinc(II) heterometallic [MFe₃S₄] ferredoxins. The trend in reduction potentials for the variants containing the [AgFe₃S₄] cluster is wild type ≤ D14C < D14H and shows the same trend as reported for the variants containing the [Fe₃S₄] cluster, but is different from the D14C < D14H < wild type trend reported for the [Fe₄S₄] ferredoxin. The similarity in the reduction potential trend for the variants containing the heterometallic [AgFe₃S₄] cluster and the [Fe₃S₄] cluster can be rationalized in terms of the electrostatic influence of the residue 14 side chains, rather than the dissociation constant of this residue, as is the case for [Fe₄S₄] ferredoxins. The trends in reduction potentials are in line with there being no electronic coupling between the silver(I) ion and the Fe₃S₄ fragment.     

Introduction of Asian strains and low genetic variation in farmed seaweeds: indications for new management practices

Seaweed farming has a crucial role in the development of future sustainable mariculture. In the same time, spreading of introduced species or genotypes from farms may threaten local ecosystems. We analyzed a molecular marker (mitochondrial cox2-3 spacers) from cultivated and wild specimen of the widely farmed seaweeds Eucheuma and Kappaphycus, collected in Zanzibar on the African east coast where commercial farming was introduced in 1989. Genotypes of presumed Asian origin were found growing on coral reefs and drifting in seagrass meadows, indicating that genotypes introduced for farming have established successfully in the wild in Zanzibar. Only a very low number of genotypes, all of Asian origin, were found in the farms. This indicates a low accessible gene pool, which can limit the capacity for adaptation to changed conditions and disease resistance in the farming system. African genotypes were found in a few sites, showing the potential for future farming of native strains. The ecological effects of the Asian genotypes introduced to coral reefs should also be further investigated in order to evaluate the risk connected with further introductions of new foreign strains.     

Core-satellite populations and seasonality of water meter biofilms in a metropolitan drinking water distribution system

Drinking water distribution systems (DWDSs) harbor the microorganisms in biofilms and suspended communities, yet the diversity and spatiotemporal distribution have been studied mainly in the suspended communities. This study examined the diversity of biofilms in an urban DWDS, its relationship with suspended communities and its dynamics. The studied DWDS in Urbana, Illinois received conventionally treated and disinfected water sourced from the groundwater. Over a 2-year span, biomass were sampled from household water meters (n=213) and tap water (n=20) to represent biofilm and suspended communities, respectively. A positive correlation between operational taxonomic unit (OTU) abundance and occupancy was observed. Examined under a ‘core-satellite'' model, the biofilm community comprised 31 core populations that encompassed 76.7% of total 16 S rRNA gene pyrosequences. The biofilm communities shared with the suspended community highly abundant and prevalent OTUs, which related to methano-/methylotrophs (i.e., Methylophilaceae and Methylococcaceae) and aerobic heterotrophs (Sphingomonadaceae and Comamonadaceae), yet differed by specific core populations and lower diversity and evenness. Multivariate tests indicated seasonality as the main contributor to community structure variation. This pattern was resilient to annual change and correlated to the cyclic fluctuations of core populations. The findings of a distinctive biofilm community assemblage and methano-/methyltrophic primary production provide critical insights for developing more targeted water quality monitoring programs and treatment strategies for groundwater-sourced drinking water systems.     

Mechanisms Preserving Insulin Action during High Dietary Fat Intake

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Statistical epistasis is a generic feature of gene regulatory networks

Functional dependencies between genes are a defining characteristic of gene networks underlying quantitative traits. However, recent studies show that the proportion of the genetic variation that can be attributed to statistical epistasis varies from almost zero to very high. It is thus of fundamental as well as instrumental importance to better understand whether different functional dependency patterns among polymorphic genes give rise to distinct statistical interaction patterns or not. Here we address this issue by combining a quantitative genetic model approach with genotype-phenotype models capable of translating allelic variation and regulatory principles into phenotypic variation at the level of gene expression. We show that gene regulatory networks with and without feedback motifs can exhibit a wide range of possible statistical genetic architectures with regard to both type of effect explaining phenotypic variance and number of apparent loci underlying the observed phenotypic effect. Although all motifs are capable of harboring significant interactions, positive feedback gives rise to higher amounts and more types of statistical epistasis. The results also suggest that the inclusion of statistical interaction terms in genetic models will increase the chance to detect additional QTL as well as functional dependencies between genetic loci over a broad range of regulatory regimes. This article illustrates how statistical genetic methods can fruitfully be combined with nonlinear systems dynamics to elucidate biological issues beyond reach of each methodology in isolation.     

Crystallographic and spectroscopic studies of peroxide-derived myoglobin compound II and occurrence of protonated FeIV O

High resolution crystal structures of myoglobin in the pH range 5.2-8.7 have been used as models for the peroxide-derived compound II intermediates in heme peroxidases and oxygenases. The observed Fe-O bond length (1.86-1.90 A) is consistent with that of a single bond. The compound II state of myoglobin in crystals was controlled by single-crystal microspectrophotometry before and after synchrotron data collection. We observe some radiation-induced changes in both compound II (resulting in intermediate H) and in the resting ferric state of myoglobin. These radiation-induced states are quite unstable, and compound II and ferric myoglobin are immediately regenerated through a short heating above the glass transition temperature (<1 s) of the crystals. It is unclear how this influences our compound II structures compared with the unaffected compound II, but some crystallographic data suggest that the influence on the Fe-O bond distance is minimal. Based on our crystallographic and spectroscopic data we suggest that for myoglobin the compound II intermediate consists of an Fe(IV)-O species with a single bond. The presence of Fe(IV) is indicated by a small isomer shift of delta = 0.07 mm/s from M?ssbauer spectroscopy. Earlier quantum refinements (crystallographic refinement where the molecular-mechanics potential is replaced by a quantum chemical calculation) and density functional theory calculations suggest that this intermediate H species is protonated.     

The essential Drosophila ATP-binding cassette domain protein, pixie, binds the 40 S ribosome in an ATP-dependent manner and is required for translation initiation

The Drosophila gene, pixie, is an essential gene required for normal growth and translation. Pixie is the fly ortholog of human RLI, which was first identified as an RNase L inhibitor, and yeast Rli1p, which has recently been shown to play a role in translation initiation and ribosome biogenesis. These proteins are all soluble ATP-binding cassette proteins with two N-terminal iron-sulfur clusters. Here we demonstrate that Pixie can be isolated from cells in complex with eukaryotic translation initiation factor 3 and ribosomal proteins of the small subunit. In addition, our analysis of polysome profiles reveals that double-stranded RNA interference-mediated depletion of Pixie results in an increase in empty 80 S ribosomes and a corresponding decrease in polysomes. Thus Pixie is required for normal levels of translation initiation. We also find that Pixie associates with the 40 S subunit on sucrose density gradients in an ATP-dependent manner. Our observations are consistent with Pixie playing a catalytic role in the assembly of complexes required for translation initiation. Thus, the function of this soluble ATP-binding cassette domain protein family in translation initiation has been conserved from yeast through to higher eukaryotes.