Localization of NADPH-protochlorophyllide reductase in plastids of barley at different greening stages

The localization of protochorophyllide (Pchlide) and of NADPH-protochlorophyllide oxidoreductase (POR, EC 1.6.99.1) within (etio)chloroplasts has been investigated at selected stages of greening of barley seedlings. Pchlide pigment and POR protein contents were evaluated in different plastid membrane fractions by fluorescence spectroscopy and immunoblot analysis using a monospecific polyclonal antibody raised against the purified enzyme. Fluorescence analysis showed the presence of Pchlide in both the envelope and thylakoid membranes. During greening, the Pchlide content, expressed on a total protein basis, decreased in thylakoid membranes, whereas it increased in the envelope membranes. POR proteins were detected mainly in thylakoid membranes at early greening stages. In contrast, the weak amount of POR proteins was associated more specifically with envelope membranes of mature chloroplasts. Whatever the greening stage, thylakoid-bound Pchlide and POR proteins were more abundant in the thylakoid regions which remained unsolubilized after mild Triton treatment used as standard procedure to prepare PS II particles. This suggests the preferential association of Pchlide and POR to the appressed regions of thylakoids. This revised version was published online in June 2006 with corrections to the Cover Date.     

Anti-stress activity of Propionibacterium freudenreichii: identification of a reactivative protein

Propionibacterium freudenreichii subsp. shermanii is known to prevent mutations caused by various agents such as N-methyl-N'-nitro-N-nitrosoguanidine, 9-aminoacridine, 4-nitro-quinoline-1-oxide and by UV radiation in both prokaryotic and eukaryotic cells. It was also shown to prevent or repair damage caused by H(2)O(2) or UV radiation in Salmonella typhimurium and Escherichia coli, a characteristic previously designated as reactivative effect. In order to characterise this effect at the molecular level, we have purified the active component from a P. freudenreichii cell-free extract using a combination of ammonium sulfate precipitation, anion-exchange and size-exclusion chromatography. The isolated 35 kDa protein was then identified using both N-terminal and internal peptide sequencing as a cysteine synthase. The latter was localised in the P. freudenreichii proteomic map. It is constitutively expressed but also clearly induced during adaptation to detergent and heat, but not acid, stresses. The biological meaning of cysteine synthase in the context of adaptation to oxidative and non-oxidative stresses is discussed.     

Cultivating the uncultured: limits, advances and future challenges

Since the invention of the Petri dish, there have been continuous efforts to improve efficiency in microbial cultivation. These efforts were devoted to the attainment for diverse growth conditions, simulation of in situ conditions and achievement of high-throughput rates. As a result, prokaryotes catalysing novel redox reactions as well as representatives of abundant, but not-yet cultured taxa, were isolated. Significant insights into microbial physiology have been made by studying the small number of prokaryotes already cultured. However, despite these numerous breakthroughs, microbial cultivation is still a low-throughput process. The main hindrance to cultivation is likely due to the prevailing lack of knowledge on targeted species. In this review, we focus on the limiting factors surrounding cultivation. We discuss several cultivation obstacles, including the loss of microbial cell–cell communication following species isolation. Future research directions, including the refinement of culture media, strategies based on cell–cell communication and high-throughput innovations, are reviewed. We further propose that a combination of these approaches is urgently required to promote cultivation of uncultured species, thereby dawning a new era in the field.     

Large-scale production of a therapeutic protein in transgenic tobacco plants: effect of subcellular targeting on quality of a recombinant dog gastric lipase

A recombinant dog gastric lipase with therapeutic potential for the treatment of exocrine pancreatic insufficiency was expressed in transgenic tobacco plants. We targeted the protein using two different signal sequences for either vacuolar retention or secretion. In both cases, an active glycosylated recombinant protein was obtained. The recombinant enzymes and the native enzyme displayed similar properties including acid resistance and acidic optimum pH. The proteolytic maturation and the specific activity of the recombinant proteins, however, were found to be dependent on subcellular compartmentalization. Expression levels of recombinant dog gastric lipase were about 5% and 7% of acid extractable plant proteins for vacuolar retention and secretion respectively. This expression system already has allowed the production of tens of grams of purified lipase through open-field culture of transgenic tobacco plants.     

The relationship between soil seed bank,above‐ground vegetation and disturbance intensity on old‐field successional permanent plots

Questions: How does disturbance and successional age influence richness, size and composition of the soil seed bank? What is the potential contribution of the soil seed bank to the plant community composition on sites differing in their successional age or disturbance intensity? Location: Experimental Botanical Garden of Göttingen University, central Germany. Methods: Above‐ground vegetation and soil seed bank were studied on formerly arable fields in a 36‐year‐old permanent plot study with five disturbance intensities, ranging from yearly ploughing via mowing to long‐term uninterrupted succession. We compared species compositions, seed densities and functional features of the seed bank and above‐ground vegetation by using several methods in parallel. Results: The seed bank was mainly composed of early successional species typical of strongly disturbed habitats. The difference between seed bank composition and above‐ground vegetation decreased with increasing disturbance intensity. The species of greatest quantitative importance in the seed bank was the non‐native forb Solidago canadensis. Conclusions: The ability of a plant community to regenerate from the soil seed bank dramatically decreases with increasing time since abandonment (successional age) and with decreasing disturbance intensity. The present study underlines that plant species typical of grasslands and woodlands are limited by dispersal capacity, owing to low capacity for accumulation of seeds in the soil and the fact that most species do not build up persistent seed banks. Rare and target species were almost absent from the seed bank and will, after local elimination, depend on reintroduction for continuation of their presence.     

Highly Efficient Flexible Hybrid Nanocrystal‐Cu(In,Ga)Se2 (CIGS) Solar Cells

A novel scheme for hybridizing inkjet‐printed thin film Cu(In,Ga)Se₂ (CIGS) solar cells with self‐assembled clusters of nanocrystal quantum dots (NQDs), which provides a 10.9% relative enhancement of the photon conversion efficiency (PCE), is demonstrated. A non‐uniform layer of NQD aggregates is deposited between the transparent conductive oxide and a CdS/CIGS p‐n junction using low cost pulsed‐spray deposition. Hybridization significantly improves the external quantum efficiency of the hybrid devices in the absorption range of the NQDs and in the red to near‐IR parts of the spectrum. The low wavelength response enhancement is found to be induced by luminescent down‐shifting (LDS) from the NQD layer, while the increase at longer wavelengths is attributed to internal scattering from NQD aggregates. LDS is demonstrated using time‐resolved spectroscopy, and the morphology of the NQD layer is investigated in fluorescence microscopy and cross‐sectional transmission electron microscopy. The influence of the NQD dose on the PCE of the hybrid devices is investigated and an optimum value is obtained. The low costs and limited material consumptions associated with pulsed‐spray deposition make these flexible hybrid devices promising candidates to help push thin‐film photovoltaic technology towards grid parity.     

Efficient isolation method for high‐quality genomic DNA from cicada exuviae

In recent years, animal ethics issues have led researchers to explore nondestructive methods to access materials for genetic studies. Cicada exuviae are among those materials because they are cast skins that individuals left after molt and are easily collected. In this study, we aim to identify the most efficient extraction method to obtain high quantity and quality of DNA from cicada exuviae. We compared relative DNA yield and purity of six extraction protocols, including both manual protocols and available commercial kits, extracting from four different exoskeleton parts. Furthermore, amplification and sequencing of genomic DNA were evaluated in terms of availability of sequencing sequence at the expected genomic size. Both the choice of protocol and exuvia part significantly affected DNA yield and purity. Only samples that were extracted using the PowerSoil DNA Isolation kit generated gel bands of expected size as well as successful sequencing results. The failed attempts to extract DNA using other protocols could be partially explained by a low DNA yield from cicada exuviae and partly by contamination with humic acids that exist in the soil where cicada nymphs reside before emergence, as shown by spectroscopic measurements. Genomic DNA extracted from cicada exuviae could provide valuable information for species identification, allowing the investigation of genetic diversity across consecutive broods, or spatiotemporal variation among various populations. Consequently, we hope to provide a simple method to acquire pure genomic DNA applicable for multiple research purposes.     

Molecular modeling of the human multidrug resistance protein 1 (MRP1/ABCC1)

Multidrug resistance protein 1 (MRP1/ABCC1) is a 190 kDa member of the ATP-binding cassette (ABC) superfamily of transmembrane transporters that is clinically relevant for its ability to confer multidrug resistance by actively effluxing anticancer drugs. Knowledge of the atomic structure of MRP1 is needed to elucidate its transport mechanism, but only low resolution structural data are currently available. Consequently, comparative modeling has been used to generate models of human MRP1 based on the crystal structure of the ABC transporter Sav1866 from Staphylococcus aureus. In these Sav1866-based models, the arrangement of transmembrane helices differs strikingly from earlier models of MRP1 based on the structure of the bacterial lipid transporter MsbA, both with respect to packing of the twelve helices and their interactions with the nucleotide binding domains. The functional importance of Tyr³²⁴ in transmembrane helix 6 predicted to project into the substrate translocation pathway was investigated.     

Graphene Doping Induced Tunability of Nanoparticles Plasmonic Resonances

Interest in graphene has been widely increasing since its discovery in 2004. Research on graphene for plasmonic applications has also boomed due to the high potential of these systems. In this article, we discuss the possible interaction between metallic NPs and graphene monolayer. We show how the contact between metallic NPs and graphene results in graphene doping. More importantly, we experimentally put into evidence the possible modulation of the plasmonic resonance of NPs by graphene doping. Understanding and evidencing this interaction is highly important both from a fundamental point of view and for specific applications such as active plasmonic devices.

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