The Bacillus subtilis signaling protein SpoIVB defines a new family of serine peptidases

The protein SpoIVB plays a key role in signaling in the sigma (K) checkpoint of Bacillus subtilis. This regulatory mechanism coordinates late gene expression during development in this organism and we have recently shown SpoIVB to be a serine peptidase. SpoIVB signals by transiting a membrane, undergoing self-cleavage, and then by an unknown mechanism activating a zinc metalloprotease, SpoIVFB, which cleaves pro-sigma (K) to its active form, sigma (K), in the outer mother cell chamber of the developing cell. In this work we have characterized the serine peptidase domain of SpoIVB. Alignment of SpoIVB with homologues from other spore formers has allowed site-specific mutagenesis of all potential active site residues within the peptidase domain. We have defined the putative catalytic domain of the SpoIVB serine peptidase as a 160-amino-acid residue segment at the carboxyl terminus of the protein. His236 and Ser378 are the most important residues for proteolysis, with Asp363 being the most probable third member of the catalytic triad. In addition, we have shown that mutations at residues Asn290 and His394 lead to delayed signaling in the sigma (K) checkpoint. The active site residues suggest that SpoIVB and its homologues from other spore formers are members of a new family of serine peptidases of the trypsin superfamily

Generic dialogue modeling for multi-application dialogue systems

We present a novel approach to developing interfaces for multi-application dialogue systems. The targeted interfaces allow transparent switching between a large number of applications within one system. The approach, based on the Rapid Dialogue Prototyping Methodology (RDPM) and the Vector Space model techniques from Information Retrieval, is composed of three main steps: (1) producing finalized dia logue models for applications using the RDPM, (2) designing an application interaction hierarchy, and (3) navigating between the applications based on the user's application of interest

The PDZ domain of the SpoIVB serine peptidase facilitates multiple functions

During spore formation in Bacillus subtilis, the SpoIVB protein is a critical component of the sigma (K) regulatory checkpoint. SpoIVB has been shown to be a serine peptidase that is synthesized in the spore chamber and which self-cleaves, releasing active forms. These forms can signal proteolytic processing of the transcription factor sigma (K) in the outer mother cell chamber of the sporulating cell. This forms the basis of the sigma (K) checkpoint and ensures accurate sigma (K)-controlled gene expression. SpoIVB has also been shown to activate a second distinct process, termed the second function, which is essential for the formation of heat-resistant spores. In addition to the serine peptidase domain, SpoIVB contains a PDZ domain. We have altered a number of conserved residues in the PDZ domain by site-directed mutagenesis and assayed the sporulation phenotype and signaling properties of mutant SpoIVB proteins. Our work has revealed that the SpoIVB PDZ domain could be used for up to four distinct processes, (i) targeting of itself for trans proteolysis, (11) binding to the protease inhibitor BofC, (iii) signaling of pro-sigma (K) processing, and (iv) signaling of the second function of SpoIVB

Mixture-Coupling theory: a mathematical model of coupled dual chemical osmosis and sorption

Very low permeability geomaterials have been used widely as barriers for landfill or nuclear waste disposal. Such materials can act as an actual semipermeable membrane. Most recently, chemical osmosis & thermal osmosis have been found important in such membrane porous media, but the theoretical and mathematical development remains a challenge due to the complexity of couplings of multiple phases and components, which thereby presents a significant bundle for scientific understanding and engineering application. This paper provides a mathematical model for dual chemical osmosis and chemical reaction based on mixture coupling theory, extending Darcy’s law with chemical osmosis and reaction. The modified Darcy’s law can be applied in any chemical disturbed underground zone

A discourse analysis of trainee teacher identity in online discussion forums

Teacher education involves an identity transformation for trainees from being a student to being a teacher. This discourse analysis examined the online discussion board communications of a cohort of trainee teachers to better understand the situated identities of the trainees and how they were presented online. Their discussion board posts were the primary method of communication during placement periods and, as such, provided insight into how the trainees situated their identities in terms of being a student or being a teacher. During the analysis, the community boundaries, language and culture were explored along with the tutor's power and role in the identity transformation process. This involved looking at the lexis used by the students, the use of pronouns to refer to themselves and others such as teachers and pupils, the types of messages allowed in the community and the effect of the tutor's messages on their communication. The research found that the trainees felt comfortable with teaching but did not feel like teachers during the course. Tutors and school teachers need to develop an awareness of the dual nature of trainees' identities and help promote the transition from student to teacher. In the beginning of the course, trainees should be familiarised with teacher vocabulary and practical concepts in addition to pedagogical theory. Towards the end of the course, trainee identity as teachers could be promoted through the use of authentic assessments that mirror real teacher tasks and requirements

Para-cresol production by Clostridium difficile affects microbial diversity and membrane integrity of Gram-negative bacteria

Clostridium difficile is a Gram-positive spore-forming anaerobe and a major cause of antibiotic-associated diarrhoea. Disruption of the commensal microbiota, such as through treatment with broad-spectrum antibiotics, is a critical precursor for colonisation by C. difficile and subsequent disease. Furthermore, failure of the gut microbiota to recover colonisation resistance can result in recurrence of infection. An unusual characteristic of C. difficile among gut bacteria is its ability to produce the bacteriostatic compound para-cresol (p-cresol) through fermentation of tyrosine. Here, we demonstrate that the ability of C. difficile to produce p-cresol in vitro provides a competitive advantage over gut bacteria including Escherichia coli, Klebsiella oxytoca and Bacteroides thetaiotaomicron. Metabolic profiling of competitive co-cultures revealed that acetate, alanine, butyrate, isobutyrate, p-cresol and p-hydroxyphenylacetate were the main metabolites responsible for differentiating the parent strain C. difficile (630Δerm) from a defined mutant deficient in p-cresol production. Moreover, we show that the p-cresol mutant displays a fitness defect in a mouse relapse model of C. difficile infection (CDI). Analysis of the microbiome from this mouse model of CDI demonstrates that colonisation by the p-cresol mutant results in a distinctly altered intestinal microbiota, and metabolic profile, with a greater representation of Gammaproteobacteria, including the Pseudomonales and Enterobacteriales. We demonstrate that Gammaproteobacteria are susceptible to exogenous p-cresol in vitro and that there is a clear divide between bacterial Phyla and their susceptibility to p-cresol. In general, Gram-negative species were relatively sensitive to p-cresol, whereas Gram-positive species were more tolerant. This study demonstrates that production of p-cresol by C. difficile has an effect on the viability of intestinal bacteria as well as the major metabolites produced in vitro. These observations are upheld in a mouse model of CDI, in which p-cresol production affects the biodiversity of gut microbiota and faecal metabolite profiles, suggesting that p-cresol production contributes to C. difficile survival and pathogenesis.Peer reviewedFinal Published versio

In-Space Propulsion: Connectivity to In-Space Fabrication and Repair

The connectivity between new in-space propulsion technologies and the ultimate development of an in-space fabrication and repair infrastructure are described in this Technical Memorandum. A number of advanced in-space propulsion technologies are being developed by NASA, many of which are directly relevant to the establishment of such an in-space infrastructure. These include aerocapture, advanced solar-electric propulsion, solar-thermal propulsion, advanced chemical propulsion, tethers, and solar photon sails. Other, further-term technologies have also been studied to assess their utility to the development of such an infrastructure