Structural analysis of PLD3 reveals insights into the mechanism of lysosomal 5′ exonuclease-mediated nucleic acid degradation

The phospholipase D (PLD) family is comprised of enzymes bearing phospholipase activity towards lipids or endo- and exonuclease activity towards nucleic acids. PLD3 is synthesized as a type II transmembrane protein and proteolytically cleaved in lysosomes, yielding a soluble active form. The deficiency of PLD3 leads to the slowed degradation of nucleic acids in lysosomes and chronic activation of nucleic acid-specific intracellular toll-like receptors. While the mechanism of PLD phospholipase activity has been extensively characterized, not much is known about how PLDs bind and hydrolyze nucleic acids. Here, we determined the high-resolution crystal structure of the luminal N-glycosylated domain of human PLD3 in its apo- and single-stranded DNA-bound forms. PLD3 has a typical phospholipase fold and forms homodimers with two independent catalytic centers via a newly identified dimerization interface. The structure of PLD3 in complex with an ssDNA-derived thymidine product in the catalytic center provides insights into the substrate binding mode of nucleic acids in the PLD family. Our structural data suggest a mechanism for substrate binding and nuclease activity in the PLD family and provide the structural basis to design immunomodulatory drugs targeting PLD3

TapA acts as specific chaperone in TasA filament formation by strand complementation

Studying mechanisms of bacterial biofilm generation is of vital importance to understanding bacterial cell–cell communication, multicellular cohabitation principles, and the higher resilience of microorganisms in a biofilm against antibiotics. Biofilms of the nonpathogenic, gram-positive soil bacterium Bacillus subtilis serve as a model system with biotechnological potential toward plant protection. Its major extracellular matrix protein components are TasA and TapA. The nature of TasA filaments has been of debate, and several forms, amyloidic and non-Thioflavin T-stainable have been observed. Here, we present the three-dimensional structure of TapA and uncover the mechanism of TapA-supported growth of nonamyloidic TasA filaments. By analytical ultracentrifugation and NMR, we demonstrate TapA-dependent acceleration of filament formation from solutions of folded TasA. Solid-state NMR revealed intercalation of the N-terminal TasA peptide segment into subsequent protomers to form a filament composed of β-sandwich subunits. The secondary structure around the intercalated N-terminal strand β0 is conserved between filamentous TasA and the Fim and Pap proteins, which form bacterial type I pili, demonstrating such construction principles in a gram-positive organism. Analogous to the chaperones of the chaperone-usher pathway, the role of TapA is in donating its N terminus to serve for TasA folding into an Ig domain-similar filament structure by donor-strand complementation. According to NMR and since the V-set Ig fold of TapA is already complete, its participation within a filament beyond initiation is unlikely. Intriguingly, the most conserved residues in TasA-like proteins (camelysines) of Bacillaceae are located within the protomer interface

TapA acts as specific chaperone in TasA filament formation by strand complementation

Studying mechanisms of bacterial biofilm generation is of vital importance to understanding bacterial cell–cell communication, multicellular cohabitation principles, and the higher resilience of microorganisms in a biofilm against antibiotics. Biofilms of the nonpathogenic, gram-positive soil bacterium Bacillus subtilis serve as a model system with biotechnological potential toward plant protection. Its major extracellular matrix protein components are TasA and TapA. The nature of TasA filaments has been of debate, and several forms, amyloidic and non-Thioflavin T-stainable have been observed. Here, we present the three-dimensional structure of TapA and uncover the mechanism of TapA-supported growth of nonamyloidic TasA filaments. By analytical ultracentrifugation and NMR, we demonstrate TapA-dependent acceleration of filament formation from solutions of folded TasA. Solid-state NMR revealed intercalation of the N-terminal TasA peptide segment into subsequent protomers to form a filament composed of β-sandwich subunits. The secondary structure around the intercalated N-terminal strand β0 is conserved between filamentous TasA and the Fim and Pap proteins, which form bacterial type I pili, demonstrating such construction principles in a gram-positive organism. Analogous to the chaperones of the chaperone-usher pathway, the role of TapA is in donating its N terminus to serve for TasA folding into an Ig domain-similar filament structure by donor-strand complementation. According to NMR and since the V-set Ig fold of TapA is already complete, its participation within a filament beyond initiation is unlikely. Intriguingly, the most conserved residues in TasA-like proteins (camelysines) of Bacillaceae are located within the protomer interface

Bark Thickness and Heights of the Bark Transition Area of Scots Pine

The estimation of forest biomass is gaining interest not only for calculating harvesting volumes but also for carbon storage estimation. However, bark (and carbon) compounds are not distributed equally along the stem. Particularly when looking at Scots pine, a radical change in the structure of the bark along the stem can be noted. At the bark transition area, the bark changes from thick and rough to thin and smooth. The aim of our study was (1) to analyze the height of the bark transition area where the bark structure changes and (2) to analyze the effect of cardinal direction on the bark thickness. Regression analyses and forward selection were performed including measured tree height, DBH, bark thickness, crown base height and upper and lower heights of the bark transition areas of 375 trees. While the cardinal direction had no effect on bark thickness, DBH was found to have a significant effect on the heights of the bark transition areas, with stand density and tree height having a minor additional effect. These variables can be used to estimate timber volume (without bark) with higher accuracy and to predict the carbon storage potential of forest biomass according to different tree compartments and compounds

Checking the Plausibility of Modelled Nitrate Concentrations in the Leachate on Federal State Scale in Germany

In Germany, modelled nitrate concentrations in the leachate are of great importance for the development of scenarios for the long-term achievement of the groundwater quality target according to the specific requirements of the EU Water Framework Directive as well as within the context of the recently adopted general administrative regulation for the designation of nitrate-polluted areas in Germany. For the German federal states of North Rhine-Westphalia (NRW) and Rhineland-Palatinate (RLP), an area-covering modelling of mean long-term nitrate concentrations in leachate with high spatial resolution was carried out using the model system RAUMIS-mGROWA-DENUZ. Hotspot regions with nitrate concentrations in the leachate of 50 mg NO3/L and more were identified for intensively farmed areas in the Münsterland, Lower Rhine, and Vorderpfalz. The validity of modelled values was checked using measured values from 1119 preselected monitoring stations from shallow springs and aquifers filtered near to the surface with oxidizing properties. For the land use categories of urban areas, arable land, grassland, and forest, an at least good agreement of modelled nitrate concentrations in the leachate and measured nitrate concentrations in groundwater was obtained at numerous sites. An equally good agreement was obtained for 1461 measuring stations from the area of responsibility of the Erftverband, which is a major water supplier in the Lower Rhine region. Here, discrepancies have been analyzed in detail due to profound regional knowledge on observation sites. It turned out that in most cases, accuracy limitations of input data (e.g., N balance surpluses of agriculture at the municipal level, 1:50,000 soil map) have been the reason for larger deviations between observed and modelled values. In a broader sense, the case study has shown on the one hand that the model system RAUMIS-mGROWA-DENUZ is able to reliably represent interrelationships and influencing factors that determine simulated nitrate concentrations in the leachate. On the other hand, it has been proven that observed nitrate concentrations in groundwater may provide a solid data source for checking the plausibility of modelled nitrate concentrations in leachate in cases where certain preselection criteria are applied

Estimation of the Number of People With Opioid Addiction in Germany

Background: Opioid addiction is one of the most common substance-related disorders worldwide, and morbidity and mortality due to opioid addiction place a heavy burden on society. Knowing the size of the population that is addicted to opioids is a prerequisite for the development and implementation of appropriate health-policy measures.Methods: Our estimate for Germany for 2016 is based on an enumeration of opioid-addicted persons who were entered in a registry of persons receiving substitution therapy, an enumeration of persons receiving outpatient and inpatient care for addiction without substitution therapy, an extrapolation to all addiction care facilities, and an estimation of the number of opioid-addicted persons who were not accounted for either in the substitution registry or in addiction care.Results: The overall estimate of the number of opioid-addicted persons in Germany in 2016 was 166 294 persons (lower and upper bounds: 164 794 and 167 794), including 123 988 men (122 968 to 125 007) and 42 307 women (41 826 to 42 787). The estimates for each German federal state per 1000 inhabitants ranged from 0.1 in Brandenburg to 3.0 in North Rhine-Westphalia and 5.5 in Bremen. The average value across Germany was 3.1 per 1000 inhabitants.Conclusion: Comparisons with earlier estimates suggest that the number of persons addicted to opioids in Germany has hardly changed over the past 20 years. Despite methodological limitations, this estimate can be considered highly valid. Nearly all persons who are addicted to opioids are in contact with the addiction care system.Schätzung der Anzahl von Personen mit einer Opioidabhängigkeit Hintergrund: Die Abhängigkeit von Opioiden zählt weltweit zu den häufigsten substanzbezogenen Störungen und verursacht eine erhebliche gesellschaftliche Belastung durch Morbidität und Mortalität. Den Umfang der opioidabhängigen Population zu kennen, ist eine Voraussetzung für gesundheitspolitische Maßnahmen.Methode: Die Schätzung für das Jahr 2016 basiert auf einer Zählung von Personen in Deutschland mit einer Opioidabhängigkeit, die im Substitutionsregister gemeldet waren, einer Zählung der in der ambulanten und stationären Suchthilfe gemeldeten Personen ohne Substitutionsbehandlung und einer Hochrechnung auf alle Einrichtungen sowie einer Schätzung der Anzahl von Personen mit einer Opioidabhängigkeit, die weder im Substitutionsregister noch in der Suchthilfe erfasst waren.Ergebnisse: Für Deutschland ergibt sich eine Gesamtschätzung von 166 294 Personen (untere und obere Grenzen: 164 794; 167 794) mit einer Opioidabhängigkeit, wobei 123 988 Männer (122 968; 125 007) und 42 307 Frauen (41 826; 42 787) betroffen sind. Die Schätzungen für die Bundesländer pro 1 000 Einwohner schwanken zwischen 0,1 in Brandenburg, 3,0 in Nordrhein-Westfalen bis 5,5 Bremen. Der deutschlandweite Durchschnittswert beträgt 3,1/1000 Einwohner.Schlussfolgerungen: Vergleiche mit früheren Schätzungen legen den Schluss nahe, dass sich der Umfang der Personen mit einer Opioidabhängigkeit in Deutschland in den letzten 20 Jahren kaum verändert hat. Trotz methodischer Einschränkungen ist von einer hohen Validität der Schätzung auszugehen. Nahezu alle Personen mit einer Opioidabhängigkeit sind in Kontakt mit dem Suchthilfesystem

Adhesives free bark panels: An alternative application for a waste material.

The proportion of bark in tree trunks is in the range of ~ 10-20%. This large amount of material is currently mainly considered as a by- or even waste-product by the timber processing industry. Recently, efforts towards the use of bark have been made, e.g. as a raw material to harvest different chemical compounds or as an additive for wood particle boards. Our motivation for this work was to keep the bark in an almost natural state and explore alternative processes and applications for use. The traditional method of de-barking tree trunks by peeling was used to harvest large bark pieces. Two pieces of peeled bark were placed crosswise, with the rhytidom side (outer bark) facing each other. After different conditioning steps, bark pieces were hot pressed to panels without adding adhesives. These experiments on bark samples of different Central European tree species suggest that production of panels with species dependent properties is possible and feasible. This is a step towards producing sustainable panels by using a natural waste material, while retaining its beneficial structure and its natural chemical composition