Mechanistic Insights into Cyclic Peptide Generation by DnaE Split-Inteins through Quantitative and Structural Investigation

Inteins carry out protein-splicing reactions, which are used in protein chemistry, protein engineering and biotechnological applications. Rearrangement of the order of the domains in split-inteins results in head-to-tail cyclisation of the target sequence, which can be used for genetic encoding and expression of libraries of cyclic peptides (CPs). The efficiency of the splicing reaction depends on the target sequence. Here we used mass spectrometry to assess in vivo cyclic peptide formation from different hexameric target sequences by the DnaE split-inteins from Synechocystis sp. and Nostoc punctiforme, revealing a strong impact of the target sequence and of the intein on the intracellular peptide concentration. Furthermore, we determined the crystal structures of their pre-splicing complexes, which allowed us to identify F-block Asp17 as crucial for the DnaE-mediated splicing reaction.     

Nitrogen, land and water inputs in changing cattle farming systems.: A historical comparison for France, 19th-21st centuries

This paper provides an original account of the long-term regional metabolism in relation to the cattle rearing in western France starting by the precise formulation of animal diets at three key dates of the 19th, 20th and 21st centuries. We established links between the demand in fodder of the meat and dairy sectors and the necessary inputs of nitrogen, water and land as well as the land cover changes occurring on the affected local and remote cattle acreage. The average agricultural productivity for fodder supply is estimated at about 50 kg N/ha in the mid-19th, 54 kg N/ha in the early 20th and 150 kg N/ha at the turning of the 21st century. Jointly for the dairy and meat productions, the potential efficiency in the conversion of the vegetal into animal protein more than doubled over the studied period, passing from less than 9% in the 19th to 20% in the 21st century. The current cattle sector is sustained for about 25% by land situated beyond the regional frontiers and uses water at intensities that approach or exceed the availability of renewable water. The nitrogen pollution is expressed in terms of the Net Anthropogenic Nitrogen Inputs (NANI) and, by comparison to the N recovered in products, is used to define the N-Environmental Efficiency of the farming. We discuss the historical succession of the factors that contributed to the growth of the meat and milk production and make a comparison of the impacts and policy between the local and distant resources.     

Bolus ingestion of white and green tea increases the concentration of several flavan‐3‐ols in plasma,but does not affect markers of oxidative stress in healthy non‐smokers

White tea (WT) is rich in flavan‐3‐ols as green tea (GT) and might provide health protective effects due to the strong antioxidant properties of flavan‐3‐ols. Since intervention studies with WT are lacking, we evaluated the effects of WT consumption on antioxidant status, antioxidant capacity and biomarkers of oxidative stress compared to water and GT. After an overnight fast, 70 healthy non‐smokers were randomized to consume 600 mL of WT, GT or water (control). Plasma (epi‐)catechin and epi(gallo)catechingallate, antioxidant capacity (Folin assay, trolox equivalent antioxidant capacity test), 8‐iso‐prostaglandin F_2α, ascorbic acid and uric acid were determined before and several times within 8 h after consumption. DNA strand breaks were measured in vivo and ex vivo (H₂O₂ stimulation) in leukocytes. Plasma flavan‐3‐ols significantly increased after WT and GT ingestion. Trolox equivalent antioxidant capacity was lower after 5 h in controls versus WT (p=0.031) and GT (p=0.005). Folin‐Ciocalteu reducing capacity, ascorbic and uric acid as well as markers of oxidative stress (8‐iso‐prostaglandin‐F_2α, DNA strand breaks) were not affected by the beverages. A short‐term increase of catechins does not change plasma antioxidant capacity in healthy subjects. Conclusions with respect to health protective effects of WT and GT on the basis of these biomarkers can, thus, not be drawn.     

Using prior knowledge in SVD-based parameter estimation for magnetic resonance spectroscopy--the ATP example

We introduce the knowledge-based singular value decomposition (KNOB-SVD) method for exploiting prior knowledge in magnetic resonance (MR) spectroscopy based on the SVD of the data matrix. More specifically, we assume that the MR data are well modeled by the superposition of a given number of exponentially damped sinusoidal components and that the dampings alphakappa, frequencies omegakappa, and complex amplitudes rhokappa of some components satisfy the following relations: alphakappa = alpha (alpha = unknown), omegakappa = omega + (kappa- 1)delta (omega = unknown, delta = known), and rhokappa = Ckapparho (rho = unknown, ckappa = known real constants). The adenosine triphosphate (ATP) complex, which has one triple peak and two double peaks whose dampings, frequencies, and amplitudes may in some cases be known to satisfy the above type of relations, is used as a vehicle for describing our SVD-based method throughout the paper. By means of numerical examples, we show that our method provides more accurate parameter estimates than a commonly used general-purpose SVD-based method and a previously suggested prior knowledge-based SVD method.     

Remote fluorescence imaging of dynamic concentration profiles with micrometer resolution using a coherent optical fiber bundle

Dynamic concentration profiles within the diffusion layer of an electrode were imaged in situ using fluorescence detection through a multichannel imaging fiber. In this work, a coherent optical fiber bundle is positioned orthogonal to the surface of an electrode and is used to report spatial and temporal micrometric changes in the fluorescence intensity of an initial fluorescent species. The fluorescence signal is directly related to the local concentration of a redox fluorescent reagent, which is electrochemically modulated by the electrode. Fluorescence images are collected through the optical fiber bundle during the oxidation of tris(2,2'-bipyridine)ruthenium(II) to ruthenium(III) at a diffusion-limited rate and allow the concentration profiles of Ru(II) reagent to be monitored in situ as a function of time. Tris(2,2'-bipyridine)ruthenium(II) is excited at 485 nm and emits fluorescence at 605 nm, whereas the Ru(III) oxidation state is not fluorescent. Our experiments emphasize the influence of two parameters on the micrometer spatial resolution: the numerical aperture of optical fibers within the bundle and the Ru(II) bulk concentration. The extent of the volume probed by each individual fiber of the bundle is discussed qualitatively in terms of a primary inner-filter effect and refractive index gradient. Experimentally measured fluorescence intensity profiles were found to be in very good agreement with concentration profiles predicted upon considering planar diffusion and thus validate the concept of this new application of imaging fibers. The originality of this remote approach is to provide a global view of the entire diffusion layer at a given time through one single image and to allow the time expansion of the diffusion layer to be followed quantitatively in real time.     

Loss measurements in stress-induced optical waveguides fabricated by an embossing process

Stress-induced light guides have been fabricated by an embossing process. The results of detailed loss measurements show that these waveguides can exhibit low optical attenuation.     

Flora und ?sung auf Wild?ckern der Feldflur in Niedersachsen

Zusammenfassung Um die Frage nach dem ökologischen Nutzen einjähriger Wildäcker der Feldflur zu klären, wurden auf 68 Standorten im Tief- und Bergland Niedersachsens während der Vegetationsperiode 2002 die Ansaaten und Pflanzenarten erfasst (letztere vegetationskundlich nachBraun-Blanquet). Jeder Wildacker unterliegt einer individuellen Anbaupraxis; jeder zweite (darunter 26 Stillegungsflächen) besitzt eine andere Einsaat. Das Spektrum der wildwachsenden Pflanzenarten vermittelt zwischen Ackerwildkraut-Gesellschaften und Ruderalen Beifußfluren und ist durch hohe Bauwerte Mehrjähriger gekennzeichnet. Die Artenzahl variiert von 12–63; im Mittel rund 40, im Diepholz-Nienburger Raum 24 Arten. 40% der Wildäcker beherbergen mindestens eine von 17 Arten der Roten Liste. Dabei sind individuenreiche Populationen die Ausnahme und nur auf Standorten mit langer Ackernutzung zu finden. Die quantitative Erfassung der von Rehwild und Feldhase verbissenen Pflanzenarten (Individuenzahl) sowie die Feststellung der pflanzenartspezifischen Äsungspräferenz (Anteil Äcker, wo Art beäst im Verhältnis zur Zahl, wo Art auftrat) ergab 23 Kultur- und 67 Wildpflanzenarten. Süßgräser, aromatische und niederliegende Pflanzenarten werden weniger angenommen; hochwüchsige, milchsaftführende und saftige bevorzugt, vor allem die Rauhe Gänsedistel (Sonchus asper L.ssp. asper). Je artenreicher der Wildacker, desto mehr Arten beäst das Wild. Die wildwachsende Flora der Wildäcker erfüllt damit eine obligate Funktion im sommerlichen Nahrungsspektrum. Eine Deckung von Kulturpflanzen über 80% führt dagegen zur Abnahme der Äsung sowie tendenziell auch zum Rückgang von Roten-Liste-Arten. Durch die regelmäßige Bestellung ackerfähiger Grenzertragsstandorte mit Feldfrüchten tragen Wildäcker zum Schutz erhaltenswerter Segetalvegetation bei, deren Restitution in Fällen kleiner Populationsgrößen der beteiligten Arten dringend geboten ist. Zur Ausschöpfung des natürlichen Standortpotentials wird ein Pilotprojekt Ausgleichsmaßnahme Wildäsungsfläche angeregt.Eingesetzt wurde ein Druckkostenzuschuß des Hessischen Ministeriums für Umwelt, Ländlichen Raum und Verbraucherschutz, für dessen Gewährung verbindlich gedankt wird. - Die Schriftleitung