Recombinant expression and purification of the 2,5-diketocamphane 1,2-monooxygenase from the camphor metabolizing Pseudomonas putida strain NCIMB 10007

Three different Baeyer-Villiger monooxygenases (BVMOs) were reported to be involved in the camphor metabolism by Pseudomonas putida NCIMB 10007. During (+)-camphor degradation, 2,5-diketocamphane is formed serving as substrate for the 2,5-diketocamphane 1,2-monooxygenase. This enzyme is encoded on the CAM plasmid and depends on the cofactors FMN and NADH and hence belongs to the group of type II BVMOs. We have cloned and recombinantly expressed the oxygenating subunit of the 2,5-diketocamphane 1,2-monooxygenase (2,5-DKCMO) in E. coli followed by His-tag-based affinity purification. A range of compounds representing different BVMO substrate classes were then investigated, but only bicyclic ketones were converted by 2,5-DKCMO used as crude cell extract or after purification. Interestingly, also (-)-camphor was oxidized, but conversion was about 3-fold lower compared to (+)-camphor. Moreover, activity of purified 2,5-DKCMO was observed in the absence of an NADH-dehydrogenase subunit

Needs, Barriers, and Support Systems for Refugee Students in Germany

This paper details the results of a study of 25 students of refugee background (SoRB) from Syria, Iran, Iraq, and Afghanistan at two technical universities of applied sciences in Berlin who were participating in preparatory courses to matriculate or were already engaging in their first semesters of study. The research shares what these refugee students feel they need in order to succeed in German higher education, and what they see as barriers. The data are organized around Baker, Ramsay, Irwin and Mile’s (2017) analytical framework of hot (familiar-informal), warm (familiar-formal) and cold (unfamiliar-formal) sources of support. This study is relevant at a time of both a massification of participation in higher education leading to greater student diversification and the development of more holistic support for all students, and an increasing mood of political agitation in countries traditionally open to migration

Deciphering the Transcriptomic Heterogeneity of Duodenal Coeliac Disease Biopsies

Coeliac disease (CD) is a clinically heterogeneous autoimmune disease with variable presentation and progression triggered by gluten intake. Molecular or genetic factors contribute to disease heterogeneity, but the reasons for different outcomes are poorly understood. Transcriptome studies of tissue biopsies from CD patients are scarce. Here, we present a high-resolution analysis of the transcriptomes extracted from duodenal biopsies of 24 children and adolescents with active CD and 21 individuals without CD but with intestinal afflictions as controls. The transcriptomes of CD patients divide into three groups—a mixed group presenting the control cases, and CD-low and CD-high groups referring to lower and higher levels of CD severity. Persistence of symptoms was weakly associated with subgroup, but the highest marsh stages were present in subgroup CD-high, together with the highest cell cycle rates as an indicator of virtually complete villous atrophy. Considerable variation in inflammation-level between subgroups was further deciphered into immune cell types using cell type de-convolution. Self-organizing maps portrayal was applied to provide high-resolution landscapes of the CD-transcriptome. We find asymmetric patterns of miRNA and long non-coding RNA and discuss the effect of epigenetic regulation. Expression of genes involved in interferon gamma signaling represent suitable markers to distinguish CD from non-CD cases. Multiple pathways overlay in CD biopsies in different ways, giving rise to heterogeneous transcriptional patterns, which potentially provide information about etiology and the course of the disease

The impact of altitude on the sleep of young elite soccer players (isa3600)

Background Altitude training is used by elite athletes to improve sports performance, but it may also disrupt sleep. The aim of this study was to examine the effects of two weeks at high altitude on the sleep of young elite athletes. Methods Participants (n=10) were members of the Australian under-17 soccer team on an 18-day (19-night) training camp in Bolivia, with 6 nights at near sea level in Santa Cruz (430 m) and 13 nights at high altitude in La Paz (3,600 m). Sleep was monitored using polysomnography during a baseline night at 430 m and three nights at 3,600 m (immediately after ascent, one week after ascent, two weeks after ascent). Data were analysed using effect size statistics. Results All results are reported as comparisons with baseline. Rapid eye movement (REM) sleep was likely lower immediately upon ascent to altitude, possibly lower after one week, and similar after two weeks. On all three nights at altitude, hypopneas and desaturations were almost certainly higher; oxygen saturation was almost certainly lower; and central apneas, respiratory arousals, and periodic breathing were very likely higher. The effects on REM sleep were common to all but one participant, but the effects on breathing were specific to only half the participants. Conclusions The immediate effects of terrestrial altitude of 3,600 m are to reduce the amount of REM sleep obtained by young elite athletes, and to cause 50% of them to have impaired breathing during sleep. REM sleep returns to normal after two weeks at altitude, but impaired breathing does not improve

The sleep of elite athletes at sea level and high altitude: A comparison of sea-level natives and high-altitude natives (ISA3600)

Background Altitude exposure causes acute sleep disruption in non-athletes, but little is known about its effects in elite athletes. The aim of this study was to examine the effects of altitude on two groups of elite athletes, that is, sea-level natives and high-altitude natives. Methods Sea-level natives were members of the Australian under-17 soccer team (n=14). High-altitude natives were members of a Bolivian under-20 club team (n=12). Teams participated in an 18-day (19 nights) training camp in Bolivia, with 6 nights at near sea level in Santa Cruz (430 m) and 13 nights at high altitude in La Paz (3600 m). Sleep was assessed on every day/night using activity monitors. Results The Australians’ sleep was shorter, and of poorer quality, on the first night at altitude compared with sea level. Sleep quality returned to normal by the end of the first week at altitude, but sleep quantity had still not stabilised at its normal level after 2 weeks. The quantity and quality of sleep obtained by the Bolivians was similar, or greater, on all nights at altitude compared with sea level. The Australians tended to obtain more sleep than the Bolivians at sea level and altitude, but the quality of the Bolivians’ sleep tended to be better than that of the Australians at altitude. Conclusions Exposure to high altitude causes acute and chronic disruption to the sleep of elite athletes who are sea-level natives, but it does not affect the sleep of elite athletes who are high-altitude natives

Wellness, fatigue and physical performance acclimatisation to a 2-week soccer camp at 3600 m (ISA3600)

Objectives To examine the time course of wellness, fatigue and performance during an altitude training camp (La Paz, 3600 m) in two groups of either sea-level (Australian) or altitude (Bolivian) native young soccer players. Methods Wellness and fatigue were assessed using questionnaires and resting heart rate (HR) and HR variability. Physical performance was assessed using HR responses to a submaximal run, a Yo-Yo Intermittent recovery test level 1 (Yo-YoIR1) and a 20 m sprint. Most measures were performed daily, with the exception of Yo-YoIR1 and 20 m sprints, which were performed near sea level and on days 3 and 10 at altitude. Results Compared with near sea level, Australians had moderate-to-large impairments in wellness and Yo-YoIR1 relative to the Bolivians on arrival at altitude. The acclimatisation of most measures to altitude was substantially slower in Australians than Bolivians, with only Bolivians reaching near sea-level baseline high-intensity running by the end of the camp. Both teams had moderately impaired 20 m sprinting at the end of the camp. Exercise HR had large associations (r>0.5–0.7) with changes in Yo-YoIR1 in both groups. Conclusions Despite partial physiological and perceptual acclimatisation, 2 weeks is insufficient for restoration of physical performance in young sea-level native soccer players. Because of the possible decrement in 20 m sprint time, a greater emphasis on speed training may be required during and after altitude training. The specific time course of restoration for each variable suggests that they measure different aspects of acclimatisation to 3600 m; they should therefore be used in combination to assess adaptation to altitude

dolls/puppets like mensch – dolls/puppets as artificial beings. Part 1.2

Die dritte Ausgabe der Zeitschrift denkste: puppe / just a bit of: doll (de:do), ein multi-disziplinäres Online-Journal für Mensch-Puppen-Diskurse, erscheint als Doppelheft, dessen gemeinsamer Themenschwerpunkt lautet: puppen/dolls like mensch – puppen als künstliche meschen. Mit diesem Fokus wird ein Thema aufgegriffen, das Menschen seit der Antike berührt und bis heute ihren Verstand und ihre Imagination, ihre Bedürfnisse und ihre Gefühle in Unruhe versetzt. In Mythologien, literarischen Fiktionen und Narrativen für Erwachsene und Kinder, in Werken der bildenden Künste, im Film, in mechanisch-technischen Anwendungen und Utopien, in den performativen Künsten, in der (Spiel-)Pädagogik und in den verschiedenen Bereichen der Popkultur wirft das Motiv der Puppe mit seinen unterschiedlichsten Ausdrucksformen immer auch existenzielle Fragen auf: Wer und was ist der Mensch? Die Puppe als künstlicher Mensch ist in gewisser Weise wie mensch, ohne Mensch zu sein. Als von Menschen geschaffene Abbilder, Vorbilder, Nachahmungen und Entwürfe des Menschen spiegeln und bestätigen Puppen vorhandene Lebenswelten und loten gleichzeitig Potenziale und Abgründe des Mensch-Seins zwischen Utopie und Dystopie, zwischen Neugier und Hingabe, zwischen Horror und Glückseligkeit, zwischen Macht und Ohnmacht aus. Puppen/dolls like mensch – der doppelte Wortsinn betont die gegebene Ambiguität der Puppen und die spannenden, ihnen innewohnenden Ambivalenzen. Im ersten Teilband (1.1) wird den Spuren und Erscheinungsformen des Puppenmotivs und der Puppe(n) – als literarisches Narrativ, als künstlerisches Motiv, als materialisiertes Objekt – vor allem im Kontext von bildender Kunst, Literatur, Fotografie, Theater und Androidentechnologien nachgegangen. Im zweiten Teilband (1.2) werden zum einen kinderliterarische und (spiel-)didaktische Texte akzentuiert, zum anderen sind hier verschiedene mediale und popkulturelle Formate aus den Bereichen Computerspiel, Comic-Film-Adaptation, Film (unterschiedlicher Genres) und dem Figurentheater versammelt sowie Thematisierungen der Verknüpfung von materiellen Artefakten und literarischen Narrativen. Rezensionen in Form von Essays über literarische Puppen-Narrative, eine Foto-Ausstellung und ein Ballett runden beide Ausgaben ab. Die zeitliche Spanne reicht vom Mittelalter bis in die Gegenwart und Zukunft und zeigt einmal mehr, wie über das Narrativ der Puppe uralte Menschheitsfragen in Traditionslinien eingebunden werden und sie auf faszinierende Weisen fortschreiben.The third edition of the journal denkste: puppe / just a bit of: doll (de: do), a multidisciplinary online journal for human-doll discourses, is a double issue whose shared thematic focus is: puppen/dolls like mensch – dolls/puppets as artificial beings. With this focus, we take up a topic that has concerned mankind since ancient times and has always upset their 'minds' and 'hearts’, their needs and feelings. In mythologies, literary fictions and narratives for adults as well as for children, in works of the visual arts, in film, in mechanical-technical applications and utopias, in the performative arts, in (play-)pedagogy and in the various fields of pop culture, the motif of the doll with its various forms of expression always raises existential questions: Who is man, what is human? The doll as an artificial human being is in a certain way like mensch without being human. As man-made images, as models, imitations and designs of humans, dolls/puppets reflect and confirm existing worlds and at the same time sound out the potentials and abysses of being human between utopia and dystopia, between curiosity and devotion, between horror and bliss, between power and powerlessness. Dolls/puppets like mensch – the double meaning of these words emphasizes the given ambiguity of the dolls/puppets and the intriguing ambivalences inherent in them. In the first part of volume (1.1) the traces and manifestations of the doll motif and of doll(s) – as literary narrative, as artistic motif, as materialized object – will be explored primarily in the context of the fine arts, of literature, photography, theater and android technologies. In the second part of the volume (1.2), on the one hand, children's literature and (play)didactic texts are accentuated; on the other hand, various media and pop-cultural formats from the fields of computer games, comic-film adaptations, films (of different genres) and puppet theater performances are gathered here, as well as issues that link material artifacts and literary narratives. Reviews in the form of essays on literary doll narratives, a photo exhibition and a ballet round off both editions. The time span extends from the Middle Ages to the present and future and shows once again how age-old questions regarding mankind and humanity are integrated into traditional lines and are carried on continuously in fascinating ways

Soccer activity profile of altitude versus sea-level natives during acclimatisation to 3600 m (ISA3600)

Objectives We investigated the effect of high altitude on the match activity profile of elite youth high altitude and sea level residents.Methods Twenty Sea Level (Australian) and 19 Altitude-resident (Bolivian) soccer players played five games, two near sea level (430 m) and three in La Paz (3600 m). Match activity profile was quantified via global positioning system with the peak 5 min period for distance ((D5peak)) and high velocity running (>4.17 m/s, HIVR5peak); as well as the 5 min period immediately subsequent to the peak for both distance (D5sub) and high-velocity running (HIVR5sub) identified using a rolling 5 min epoch. The games at 3600 m were compared with the average of the two near sea-level games.Results The total distance per minute was reduced by a small magnitude in the first match at altitude in both teams, without any change in low-velocity running. There were variable changes in HiVR, D5peak and HiVR5peak from match to match for each team. There were within-team reductions in D5peak in each game at altitude compared with those at near sea level, and this reduction was greater by a small magnitude in Australians than Bolivians in game 4. The effect of altitude on HiVR5peak was moderately lower in Australians compared with Bolivians in game 3. There was no clear difference in the effect of altitude on maximal accelerations between teams.Conclusions High altitude reduces the distance covered by elite youth soccer players during matches. Neither 13 days of acclimatisation nor lifelong residence at high altitude protects against detrimental effects of altitude on match activity profile

The Metagenome-Derived Enzymes LipS and LipT Increase the Diversity of Known Lipases

Triacylglycerol lipases (EC 3.1.1.3) catalyze both hydrolysis and synthesis reactions with a broad spectrum of substrates rendering them especially suitable for many biotechnological applications. Most lipases used today originate from mesophilic organisms and are susceptible to thermal denaturation whereas only few possess high thermotolerance. Here, we report on the identification and characterization of two novel thermostable bacterial lipases identified by functional metagenomic screenings. Metagenomic libraries were constructed from enrichment cultures maintained at 65 to 75°C and screened resulting in the identification of initially 10 clones with lipolytic activities. Subsequently, two ORFs were identified encoding lipases, LipS and LipT. Comparative sequence analyses suggested that both enzymes are members of novel lipase families. LipS is a 30.2 kDa protein and revealed a half-life of 48 h at 70°C. The lipT gene encoded for a multimeric enzyme with a half-life of 3 h at 70°C. LipS had an optimum temperature at 70°C and LipT at 75°C. Both enzymes catalyzed hydrolysis of long-chain (C12 and C14) fatty acid esters and additionally hydrolyzed a number of industry-relevant substrates. LipS was highly specific for (R)-ibuprofen-phenyl ester with an enantiomeric excess (ee) of 99%. Furthermore, LipS was able to synthesize 1-propyl laurate and 1-tetradecyl myristate at 70°C with rates similar to those of the lipase CalB from Candida antarctica. LipS represents the first example of a thermostable metagenome-derived lipase with significant synthesis activities. Its X-ray structure was solved with a resolution of 1.99 Å revealing an unusually compact lid structure

Antithetical NFATc1–Sox2 and p53–miR200 signaling networks govern pancreatic cancer cell plasticity

In adaptation to oncogenic signals, pancreatic ductal adenocarcinoma (PDAC) cells undergo epithelial-mesenchymal transition (EMT), a process combining tumor cell dedifferentiation with acquisition of stemness features. However, the mechanisms linking oncogene-induced signaling pathways with EMT and stemness remain largely elusive. Here, we uncover the inflammation-induced transcription factor NFATc1 as a central regulator of pancreatic cancer cell plasticity. In particular, we show that NFATc1 drives EMT reprogramming and maintains pancreatic cancer cells in a stem cell-like state through Sox2-dependent transcription of EMT and stemness factors. Intriguingly, NFATc1-Sox2 complex-mediated PDAC dedifferentiation and progression is opposed by antithetical p53-miR200c signaling, and inactivation of the tumor suppressor pathway is essential for tumor dedifferentiation and dissemination both in genetically engineered mouse models (GEMM) and human PDAC. Based on these findings, we propose the existence of a hierarchical signaling network regulating PDAC cell plasticity and suggest that the molecular decision between epithelial cell preservation and conversion into a dedifferentiated cancer stem cell-like phenotype depends on opposing levels of p53 and NFATc1 signaling activities