Cloning,over expression and functional attributes of serine proteases from Oceanobacillus iheyensis O.M.A18 and Haloalkaliphilic bacterium O.M.E12

Cloning, over-expression, characterization and structural and functional analysis of two alkaline proteases from the newly isolated haloalkaliphilic bacteria: Oceanobacillus iheyensis O.M.A₁8 and Haloalkaliphilic bacterium O.M.E₁2 were carried out. The cloned protease genes were over-expressed in Escherichia coli within 6 h of the IPTG induction. The protease genes were sequenced and the sequence submitted to the GenBank with the accession numbers, HM219179 and HM219182. The recombinant proteases were active in the range of pH 8–11 and temperature 30–50 °C. The amino acid sequences of the alkaline proteases displayed hydrophobic character and stable configurations. The amino acids Asp 141, His 171 and Ser 324 formed the catalytic triad, while Ile, Leu and Ser were other amino acid moieties present in the active site. The characteristics of the recombinant proteases were compared and found to be similar to their native counterparts. On the basis of the in-silico analysis and inhibitor studies, the enzymes were confirmed as serine proteases. The study hold significance as only limited enzymes from the haloalkaliphilic bacteria have been cloned, sequenced and analyzed for the structure and function analysis.     

Applications of Hydrolytic and Decarboxylating Enzymes in Biotransformations

Peptides, and oligosaccharides and glycosides, can be synthesised by making use of the ‘reverse hydrolytic activity’ of proteases and glycosidases respectively. In applying these enzymes to the practical synthesis of these classes of compound, several factors need to be considered, namely the need to shift the rate-determining step through the use of activated substrates, the need to minimise competing hydrolysis of these and the need to minimise hydrolysis of the products. In spite of these problems, the enzymatic methods have many attractive features, not least amongst which is the absolute control of stereochemistry in acyl transfer and glycosyl transfer respectively.     

生态系统服务簇空间演变轨迹及其社会-生态驱动的地理探测——以大连市为例

生态系统服务簇是多种生态系统服务的组合，明晰生态系统服务簇及其自然-社会经济驱动因素，对生态系统服务内部相互依赖机制识别、实现多种生态系统服务间良性互动具有重要意义。目前生态系统服务簇的识别已得到广泛应用，但对多种生态系统服务之间交互作用的动态过程与影响机理的认识还不够深刻。针对目前多种生态系统服务间交互作用动态演化分析及其社会-生态驱动机理研究不足的现状，以我国北方沿海重要中心城市——大连市为例，选取食物供给、水源涵养、固碳释氧、土壤保持、生境质量和景观美学6种关键服务。采用Spearman相关性分析方法探究生态系统服务权衡与协同关系，借助自组织网络方法识别生态系统服务簇，进一步分析多种生态系统服务间交互作用的时空分异特征，运用地理探测器探究其空间分异影响因素。结果表明：(1)食物供给与土壤保持存在极显著的权衡关系■,土壤保持与景观美学存在极显著的协同关系■。(2)2005—2015年大连市生态保育簇空间格局较稳定，水源涵养簇、食物供给簇与服务枯竭簇之间轨迹变化明显，城市扩张与服务枯竭簇演变具有一致性。(3)高程、归一化植被指数是影响生态系统服务簇空间分布的关键自然因素，而土地利用强...     

Forest floor photosynthesis and respiration in a drained peatland forest in southern Finland

Background: Although forest floor forms a large biomass pool in forested peatlands, little is known about its role in ecosystem carbon (C) dynamics.

Aim: We aimed to quantify forest floor photosynthesis (P _FF) and respiration (R _FF) as a part of overall C dynamics in a drained peatland forest in southern Finland.

Methods: We measured net forest floor CO₂ exchange with closed chambers and reconstructed seasonal CO₂ exchange in the prevailing plant communities.

Results: The vegetation was a mosaic of plant communities that differed in CO₂ exchange dynamics. The reconstructed growing season P _FF was highest in the Sphagnum community and lowest in the feather moss communities. On the contrary, R _FF was highest in the feather moss communities and lowest in the Sphagnum community. CO₂ assimilated by the forest floor was 20–30% of the total CO₂ assimilated by the forest. The forest floor was a net CO₂ source to the atmosphere, because respiration from ground vegetation, tree roots and decomposition of soil organic matter exceeded the photosynthesis of ground vegetation.

Conclusions: Tree stand dominates C fluxes in drained peatland forests. However, forest floor vegetation can have a noticeable role in the C cycle of peatlands drained for forestry. Similarly to natural mires, Sphagnum moss-dominated communities were the most efficient assimilators of C.     

Defect of Fe-S cluster binding by DNA polymerase δ in yeast suppresses UV-induced mutagenesis,but enhances DNA polymerase ζ – dependent spontaneous mutagenesis

Eukaryotic genomes are duplicated by a complex machinery, utilizing high fidelity replicative B-family DNA polymerases (pols) α, δ and ε. Specialized error-prone pol ζ, the fourth B-family member, is recruited when DNA synthesis by the accurate trio is impeded by replication stress or DNA damage. The damage tolerance mechanism dependent on pol ζ prevents DNA/genome instability and cell death at the expense of increased mutation rates. The pol switches occurring during this specialized replication are not fully understood. The loss of pol ζ results in the absence of induced mutagenesis and suppression of spontaneous mutagenesis. Disruption of the Fe-S cluster motif that abolish the interaction of the C-terminal domain (CTD) of the catalytic subunit of pol ζ with its accessory subunits, which are shared with pol δ, leads to a similar defect in induced mutagenesis. Intriguingly, the pol3-13 mutation that affects the Fe-S cluster in the CTD of the catalytic subunit of pol δ also leads to defective induced mutagenesis, suggesting the possibility that Fe-S clusters are essential for the pol switches during replication of damaged DNA. We confirmed that yeast strains with the pol3-13 mutation are UV-sensitive and defective in UV-induced mutagenesis. However, they have increased spontaneous mutation rates. We found that this increase is dependent on functional pol ζ. In the pol3-13 mutant strain with defective pol δ, there is a sharp increase in transversions and complex mutations, which require functional pol ζ, and an increase in the occurrence of large deletions, whose size is controlled by pol ζ. Therefore, the pol3-13 mutation abrogates pol ζ-dependent induced mutagenesis, but allows for pol ζ recruitment for the generation of spontaneous mutations and prevention of larger deletions. These results reveal differential control of the two major types of pol ζ-dependent mutagenesis by the Fe-S cluster present in replicative pol δ.     

Diel vertical migration ofEudiaptomus gracilis during a short summer period

Several aspects of a diel vertical migration (DVM) of adultEudiaptomus gracilis in Lake Maarsseveen (The Netherlands) are described. The period of DVM lasted from the end of May until the middle of August. On May 21, 1989, the population was found divided into a deep dwelling part and a part in the upper five meter. Large shoals of juvenile perch were observed in the open water for the first time. On June 7, the whole population was down below 10 m and concentrated in a zone of high chlorphyll-a concentrations. One week later, a regular DVM was performed. The amplitude of this migration gradually decreased towards the end of the migration period. The ascent in the evening and the descent in the morning took place after sunset and before sunrise, respectively. The movements coincided with high relative changes in light intensity. Population size increased rapidly during the period of DVM but decreased again before the end of this period.     

The lifetime of molecular (Davydov) solitons

The lifetime of Davydov solitons in a one-dimensional system is studied theoretically. The process of thermalization and the properties of solitons at finite temperature are investigated and the processes of soliton creation and disintegration are discussed.     

Colonization and extinction of land planarians (Platyhelminthes,Tricladida) in a Brazilian Atlantic Forest regrowth remnant

Long-term assessments of species assemblages are valuable tools for detecting species ecological preferences and their dispersal tracks, as well as for assessing the possible effects of alien species on native communities. Here we report a 50-year-long study on population dynamics of the four species of land flatworms (Platyhelminthes, Tricladida, Terricola) that have colonized or become extinct in a 70-year-old Atlantic Forest regrowth remnant through the period 1955–2006. On the one hand, the two initially most abundant species, which are native to the study site, Notogynaphallia ernesti and Geoplana multicolor have declined over decades and at present do not exist in the forest remnant. The extinction of these species is most likely related with their preference for open vegetation areas, which presently do not exist in the forest remnant. On the other hand, the neotropical Geoplaninae 1 and the exotic Endeavouria septemlineata were detected in the forest only very recently. The long-term study allowed us to conclude that Geoplaninae 1 was introduced into the study area, although it is only known from the study site. Endeavouria septemlineata, an active predator of the exotic giant African snail, is originally known from Hawaii. This land flatworm species was observed repeatedly in Brazilian anthropogenic areas, and this is the first report of the species in relatively well preserved native forest, which may be evidence of an ongoing adaptive process. Monitoring of its geographic spread and its ecological role would be a good practice for preventing potential damaging effects, since it also feeds on native mollusk fauna, as we observed in lab conditions. Júlio Pedroni: Granted by CNPQ–Brazil.     

Metabolic control at the cytosol–mitochondria interface in different growth phases of CHO cells

Metabolism at the cytosol–mitochondria interface and its regulation is of major importance particularly for efficient production of biopharmaceuticals in Chinese hamster ovary (CHO) cells but also in many diseases. We used a novel systems-oriented approach combining dynamic metabolic flux analysis and determination of compartmental enzyme activities to obtain systems level information with functional, spatial and temporal resolution. Integrating these multiple levels of information, we were able to investigate the interaction of glycolysis and TCA cycle and its metabolic control. We characterized metabolic phases in CHO batch cultivation and assessed metabolic efficiency extending the concept of metabolic ratios. Comparing in situ enzyme activities including their compartmental localization with in vivo metabolic fluxes, we were able to identify limiting steps in glycolysis and TCA cycle. Our data point to a significant contribution of substrate channeling to glycolytic regulation. We show how glycolytic channeling heavily affects the availability of pyruvate for the mitochondria. Finally, we show that the activities of transaminases and anaplerotic enzymes are tailored to permit a balanced supply of pyruvate and oxaloacetate to the TCA cycle in the respective metabolic states. We demonstrate that knowledge about metabolic control can be gained by correlating in vivo metabolic flux dynamics with time and space resolved in situ enzyme activities.