739 research outputs found

    Secretion and assembly of functional mini-cellulosomes from synthetic chromosomal operons in Clostridium acetobutylicum ATCC 824.

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    Background: Consolidated bioprocessing (CBP) is reliant on the simultaneous enzyme production, saccharification of biomass, and fermentation of released sugars into valuable products such as butanol. Clostridial species that produce butanol are, however, unable to grow on crystalline cellulose. In contrast, those saccharolytic species that produce predominantly ethanol, such as Clostridium thermocellum and Clostridium cellulolyticum, degrade crystalline cellulose with high efficiency due to their possession of a multienzyme complex termed the cellulosome. This has led to studies directed at endowing butanol-producing species with the genetic potential to produce a cellulosome, albeit by localising the necessary transgenes to unstable autonomous plasmids. Here we have explored the potential of our previously described Allele-Coupled Exchange (ACE) technology for creating strains of the butanol producing species Clostridium acetobutylicum in which the genes encoding the various cellulosome components are stably integrated into the genome. Results: We used BioBrick2 (BB2) standardised parts to assemble a range of synthetic genes encoding C. thermocellum cellulosomal scaffoldin proteins (CipA variants) and glycoside hydrolases (GHs, Cel8A, Cel9B, Cel48S and Cel9K) as well as synthetic cellulosomal operons that direct the synthesis of Cel8A, Cel9B and a truncated form of CipA. All synthetic genes and operons were integrated into the C. acetobutylicum genome using the recently developed ACE technology. Heterologous protein expression levels and mini-cellulosome self-assembly were assayed by western blot and native PAGE analysis. Conclusions: We demonstrate the successful expression, secretion and self-assembly of cellulosomal subunits by the recombinant C. acetobutylicum strains, providing a platform for the construction of novel cellulosomes. © 2013 Kovács et al.; licensee BioMed Central Ltd

    The application of passive sampler (DGT) technology for improved understanding of metal behaviour at a marine disposal site

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    Metal behaviour and availability at a contaminated dredge material disposal site within UK waters has been investigated using Diffusive Gradient in Thin films (DGT) passive sampling technology. Three stations representing contrasting history and presence of maintenance dredge disposal, including a control station outside the disposal site, have been studied and depth profiles of fluxes of different metals (Fe, Mn, Pb, Cu, Cd, Cr, Ni, Zn) to the binding gel (Chelex 100) have been derived. Higher flux rates and shallower mobilisation of metals (Mn and Fe) to the binding gel were observed at the disposal stations compared to the control station. Here we describe metal mobilization at different depths, linking the remobilization of Fe2+ and Mn2+ to the sediment (re)supply of other heavy metals of interest with a focus on Cd, Ni and Pb and as they are on the Water Framework Directive (WFD) list of priority substances and OSPAR list of priority pollutants. Results showed that Cd, Pb and Ni exhibited signs of resupply at the sediment-water interface (SWI). There was a potential increased mobilisation and source to the water column of Pb and Ni at the disposal site stations, but there was no Cd source, despite higher total loadings. This information has the potential to improve our current understanding of metal cycles at disposal sites. This work can be used as an indication of likely metal bioavailability and also assist in determining whether the sites act as sources or sinks of heavy metals. This information could assist disposal site monitoring and dredge material licensing

    A Novel Extracellular Metallopeptidase Domain Shared by Animal Host-Associated Mutualistic and Pathogenic Microbes

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    The mucosal microbiota is recognised as an important factor for our health, with many disease states linked to imbalances in the normal community structure. Hence, there is considerable interest in identifying the molecular basis of human-microbe interactions. In this work we investigated the capacity of microbes to thrive on mucosal surfaces, either as mutualists, commensals or pathogens, using comparative genomics to identify co-occurring molecular traits. We identified a novel domain we named M60-like/PF13402 (new Pfam entry PF13402), which was detected mainly among proteins from animal host mucosa-associated prokaryotic and eukaryotic microbes ranging from mutualists to pathogens. Lateral gene transfers between distantly related microbes explained their shared M60-like/PF13402 domain. The novel domain is characterised by a zinc-metallopeptidase-like motif and is distantly related to known viral enhancin zinc-metallopeptidases. Signal peptides and/or cell surface anchoring features were detected in most microbial M60-like/PF13402 domain-containing proteins, indicating that these proteins target an extracellular substrate. A significant subset of these putative peptidases was further characterised by the presence of associated domains belonging to carbohydrate-binding module family 5/12, 32 and 51 and other glycan-binding domains, suggesting that these novel proteases are targeted to complex glycoproteins such as mucins. An in vitro mucinase assay demonstrated degradation of mammalian mucins by a recombinant form of an M60-like/PF13402-containing protein from the gut mutualist Bacteroides thetaiotaomicron. This study reveals that M60-like domains are peptidases targeting host glycoproteins. These peptidases likely play an important role in successful colonisation of both vertebrate mucosal surfaces and the invertebrate digestive tract by both mutualistic and pathogenic microbes. Moreover, 141 entries across various peptidase families described in the MEROPS database were also identified with carbohydrate-binding modules defining a new functional context for these glycan-binding domains and providing opportunities to engineer proteases targeting specific glycoproteins for both biomedical and industrial applications

    Transitioning in higher education: An exploration of psychological and contextual factors affecting student satisfaction

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    © 2017 UCU. In view of recent changes in the higher education sector, such as increased tuition fees, a greater focus has been placed on widening participation initiatives and monitoring student satisfaction. The aims of the current study were twofold: (1) to explore whether pre-entry programmes foster successful transition to higher education, and (2) to examine longitudinally the factors associated with course satisfaction. Eighty-eight first-year psychology students completed a questionnaire measuring academic self-efficacy, social identity and student satisfaction at the start (Time 1, November 2015) and end (Time 2, March 2016) of the academic year. Findings indicated that students who participated in a pre-entry programme reported higher academic self-efficacy and satisfaction compared to typical route students. Moreover, academic self-efficacy predicted student satisfaction at the start of the academic year, whereas in-group affect (a facet of social identity) predicted this at the end of the academic year. The current findings indicate that pre-entry programmes may have a positive impact on students’ sense of academic self-efficacy. On a more general level, the findings also suggest that academic self-efficacy and social identity may be key indicators of student satisfaction. This highlights the complexities of the concept of ‘student satisfaction’, and demonstrates the utility of examining multiple factors relating to student satisfaction across different time points

    Characterisation of anhydro-sialic acid transporters from mucosa-associated bacteria

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    Sialic acid (Sia) transporters are critical to the capacity of host-associated bacteria to utilise Sia for growth and/or cell surface modification. While N-acetyl-neuraminic acid (Neu5Ac)-specific transporters have been studied extensively, little is known on transporters dedicated to anhydro-Sia forms such as 2,7-anhydro-Neu5Ac (2,7-AN) or 2,3-dehydro-2-deoxy-Neu5Ac (Neu5Ac2en). Here, we used a Sia-transport-null strain of Escherichia coli to investigate the function of members of anhydro-Sia transporter families previously identified by computational studies. First, we showed that the transporter NanG, from the Glycoside-Pentoside-Hexuronide:cation symporter family, is a specific 2,7-AN transporter, and identified by mutagenesis a crucial functional residue within the putative substrate-binding site. We then demonstrated that NanX transporters, of the Major Facilitator Superfamily, also only transport 2,7-AN and not Neu5Ac2en nor Neu5Ac. Finally, we provided evidence that SiaX transporters, of the Sodium-Solute Symporter superfamily, are promiscuous Neu5Ac/Neu5Ac2en transporters able to acquire either substrate equally well. The characterisation of anhydro-Sia transporters expands our current understanding of prokaryotic Sia metabolism within host-associated microbial communities

    Open Science in Software Engineering

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    Open science describes the movement of making any research artefact available to the public and includes, but is not limited to, open access, open data, and open source. While open science is becoming generally accepted as a norm in other scientific disciplines, in software engineering, we are still struggling in adapting open science to the particularities of our discipline, rendering progress in our scientific community cumbersome. In this chapter, we reflect upon the essentials in open science for software engineering including what open science is, why we should engage in it, and how we should do it. We particularly draw from our experiences made as conference chairs implementing open science initiatives and as researchers actively engaging in open science to critically discuss challenges and pitfalls, and to address more advanced topics such as how and under which conditions to share preprints, what infrastructure and licence model to cover, or how do it within the limitations of different reviewing models, such as double-blind reviewing. Our hope is to help establishing a common ground and to contribute to make open science a norm also in software engineering.Comment: Camera-Ready Version of a Chapter published in the book on Contemporary Empirical Methods in Software Engineering; fixed layout issue with side-note

    Evaluating impacts of bottom trawling and hypoxia on benthic communities at the local, habitat, and regional scale using a modelling approach

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    Bottom trawling disturbance and hypoxia are affecting marine benthic habitats worldwide. We present an approach to predict their effects on benthic communities, and use the approach to estimate the state, the biomass relative to carrying capacity, of the Baltic Sea at the local, habitat, and regional scale. Responses to both pressures are expected to depend on the longevity of fauna, which is predicted from benthic data from 1558 locations. We find that communities in low-salinity regions mostly consist of short-lived species, which are, in our model, more resilient than those of the saline areas. The model predicts that in 14% of the Baltic Sea region benthic biomass is reduced by at least 50%, whereas an additional 8% of the region has reductions of 10-50%. The effects of hypoxia occur over larger spatial scales and lead to a low state of especially deep habitats. The approach is based on a simple characterization of the benthic community, which comes with high uncertainty, but allows for the identification of benthic habitats that are at greatest risk and prioritization of management actions at the regional scale. This information supports the development of sustainable approaches to manage impact of human activities on benthic ecosystems.</p
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