Crystallization of the regulatory and effector domains of the key sporulation response regulator Spo0A

The key response-regulator gene of sporulation, spo0A, has been cloned from Bacillus stearothermophilus and the encoded protein purified. The DNA-binding and phospho-acceptor domains of Spo0A have been prepared by tryptic digestion of the intact protein and subsequently crystallized in forms suitable for X-ray crystallographic studies. The DNA-binding domain has been crystallized in two forms, one of which diffracts X-rays to beyond 2.5 Angstrom spacing. The crystals of the phospho-acceptor domain diffract X-rays beyond 2.0 Angstrom spacing using synchrotron radiation

The trans-activation domain of the sporulation response regulator Spo0A revealed by X-ray crystallography

Sporulation in Bacillus involves the induction of scores of genes in a temporally and spatially co-ordinated programme of cell development. Its initiation is under the control of an expanded two-component signal transduction system termed a phosphorelay. The master control element in the decision to sporulate is the response regulator, Spo0A, which comprises a receiver or phosphoacceptor domain and an effector or transcription activation domain. The receiver domain of Spo0A shares sequence similarity with numerous response regulators, and its structure has been determined in phosphorylated and unphosphorylated forms. However, the effector domain (C-Spo0A) has no detectable sequence similarity to any other protein, and this lack of structural information is an obstacle to understanding how DNA binding and transcription activation are controlled by phosphorylation in Spo0A. Here, we report the crystal structure of C-Spo0A from Bacillus stearothermophilus revealing a single alpha -helical domain comprising six alpha -helices in an unprecedented fold. The structure contains a helix-turn-helix as part of a three alpha -helical bundle reminiscent of the catabolite gene activator protein (CAP), suggesting a mechanism for DNA binding. The residues implicated in forming the sigma (A)-activating region clearly cluster in a flexible segment of the polypeptide on the opposite side of the structure from that predicted to interact with DNA. The structural results are discussed in the context of the rich array of existing mutational data

Spo0A∼P Imposes a Temporal Gate for the Bimodal Expression of Competence in Bacillus subtilis

ComK transcriptionally controls competence for the uptake of transforming DNA in Bacillus subtilis. Only 10%–20% of the cells in a clonal population are randomly selected for competence. Because ComK activates its own promoter, cells exceeding a threshold amount of ComK trigger a positive feedback loop, transitioning to the competence ON state. The transition rate increases to a maximum during the approach to stationary phase and then decreases, with most cells remaining OFF. The average basal rate of comK transcription increases transiently, defining a window of opportunity for transitions and accounting for the heterogeneity of competent populations. We show that as the concentration of the response regulator Spo0A∼P increases during the entry to stationary phase it first induces comK promoter activity and then represses it by direct binding. Spo0A∼P activates by antagonizing the repressor, Rok. This amplifies an inherent increase in basal level comK promoter activity that takes place during the approach to stationary phase and is a general feature of core promoters, serving to couple the probability of competence transitions to growth rate. Competence transitions are thus regulated by growth rate and temporally controlled by the complex mechanisms that govern the formation of Spo0A∼P. On the level of individual cells, the fate-determining noise for competence is intrinsic to the comK promoter. This overall mechanism has been stochastically simulated and shown to be plausible. Thus, a deterministic mechanism modulates an inherently stochastic process

Przyczyny stagnacji konsolidacji gruntów w Słowacji i Republice Czeskiej

Causes of different evolution of land consolidation in the Czech Republic (CR) and Slovakia (SR) are documented and analyzed. Land fragmentation, methodical guidelines, legislative measures, financing and implementation of land consolidation projects are compared. Extensive, broad, complex land consolidation (LC) brings direct and indirect economic, environmental, social and landscape benefits for land owners and communities alike. It is a planning and development tool that is crucial for regional development. Authors focus on success of LC projects (measured by numbers of accepted projects relative to the country size) and their historical backgrounds in both neighbouring countries. Comprehensive land consolidation (CLC) and simple land consolidation (SLC) are examined. Approach to LC is similar in both countries. Demand for solving certain problems (e.g. land fragmentation, ownership fragmentation) is higher in Slovakia. Comprehensive land consolidation projects were initiated earlier in Slovakia than in the Czech Republic. But the current situation is significantly worse in Slovakia than in the Czech Republic. Risk of promoting particular interests resulting in an environment with disrupted dynamics of land consolidation is mentioned. Slovakia has a chance to change current dismal situation in the field of land consolidation only with the support of landowners.W pracy udokumentowano i przeanalizowano przyczyny odmiennego rozwoju konsolidacji gruntów w Republice Czeskiej i Słowacji. Porównano fragmentację ziemi, wskazania metodyczne, środki prawne, finansowanie i wdrażanie projektów konsolidacji gruntów. Ekstensywna, szeroka i złożona konsolidacja gruntów przynosi bezpośrednie i pośrednie korzyści ekonomiczne, środowiskowe, społeczne i krajobrazowe, zarówno właścicielom ziemi, jak i społecznościom. Jest narzędziem planowania i rozwoju istotnym dla rozwoju regionalnego. Autorzy skupili się na sukcesie projektów konsolidacji gruntów (mierzonym liczbą zaakceptowanych projektów w stosunku do powierzchni kraju) i na ich historycznym tle w obu sąsiadujących państwach. Badano całościową i prostą konsolidację gruntów. Podejście do konsolidacji jest podobne w obu krajach. W Słowacji istnieje większa potrzeba rozwiązywania pewnych problemów (np. fragmentacja ziemi, rozdrobnienie własnościowe). Projekty całościowej konsolidacji gruntów powstały wcześniej w Słowacji niż w Republice Czeskiej, ale obecna sytuacja jest znacząco gorsza w Słowacji. Wymienia się ryzyko realizacji partykularnych interesów, które skutkują przerwaniem dynamicznej konsolidacji gruntów. Słowacja ma szansę na zmianę obecnej fatalnej sytuacji jedynie w warunkach wsparcia ze strony właścicieli ziemskich

Leaching of Metals from Incineration Bottom Ash Using Organic Acid

The municipal solid waste incineration bottom ash (IBA) is increasingly used as a secondary resource for civil engineering and various other applications. IBA contains various metals. The presence of these metals may limit the use of IBA as a secondary resource. This can be overcome by removing metals from IBA. This study focused on determination of metal contents of IBA and leaching of these metals. The results show that calcium (Ca), iron (Fe), aluminum (Al), silicon (Si), and sodium (Na) are the major metals present in IBA. Many other metals ranging from magnesium (Mg) to mercury (Hg) were also present in varying quantities. In the present study, four organic acids, namely, citric, malic, oxalic, and gluconic acids have been used to compare the metal leaching from IBA. The citric acid was found to be the most effective leaching agent. Various process parameters for metal leaching were optimized. It was found that the maximum metal leaching was achieved in 48 h. The 1 M citric acid, 100 mL working volume, 150 rpm, and 30 °C were the optimal conditions to leach 100 wt% Si, Na, Au, Sb, respectively. Around 99 wt% Mg, 95 wt% K, 93 wt% Cd, 88 wt% Al, 84 wt% Ca, 80 wt% Fe, 79 wt% Sr, 77 wt% Pb, Zn, As respectively, 67 wt% Cu, V, Ag respectively, 66 wt% Mn, 64 wt% Co, 56 wt% Cr, 43 wt% Ni, Sn respectively, 40 wt% Se, 21 wt% Ba, 10 wt% Hg were leached using citric acid at optimal conditions. The leaching efficiency decreased with increasing pulp density. An optimal pulp density for metal leaching was observed at 1% (w/v). The method described in the present study provides an alternative environmentally friendly process to remove metals from IBA. This will facilitate the recycling of metal-free IBA for geotechnical applications.National Environmental Agency (NEA)The authors would like to acknowledge the financial support from the Environment Technology Research Program (ETRP), the National Environment Agency, Singapore (ETRP Nos. 1301 104)