Precursor ion scanning for detection and structural characterization of heterogeneous glycopeptide mixtures

AbstractThe structure of N-linked glycans is determined by a complex, anabolic, intracellular pathway but the exact role of individual glycans is not always clear. Characterization of carbohydrates attached to glycoproteins is essential to aid understanding of this complex area of biology. Specific mass spectral detection of glycopeptides from protein digests may be achieved by on-line HPLC-MS, with selected ion monitoring (SIM) for diagnostic product ions generated by cone voltage fragmentation, or by precursor ion scanning for terminal saccharide product ions, which can yield the same information more rapidly. When glycosylation is heterogeneous, however, these approaches can result in spectra that are complex and poorly resolved. We have developed methodology, based around precursor ion scanning for ions of high m/z, that allows site specific detection and structural characterization of glycans at high sensitivity and resolution. These methods have been developed using the standard glycoprotein, fetuin, and subsequently applied to the analysis of the N-linked glycans attached to the scrapie-associated prion protein, PrPSc. These glycans are highly heterogeneous and over 30 structures have been identified and characterized site specifically. Product ion spectra have been obtained on many glycopeptides confirming structure assignments. The glycans are highly fucosylated and carry Lewis X or sialyl Lewis X epitopes and the structures are in-line with previous results. [Abbreviations: Hex–Hexose, C6H12O6 carbohydrates, including mannnose and galactose; HexNAc—N-acetylhexosamine, C8H15NO6 carbohydrates, including N-acetylglucosamine and N-acetylgalactosamine; GlcNAc—N-acetylglucosamine; GalNAc—N-acetylgalactosamine; Fuc–Fucose; NeuAC—N-acetylneuraminic acid or sialic acid; TSE—Transmissible Spongiform Encephalopathy.

Field phenomics:will it enable crop improvement?

Field phenomics has been identified as a promising enabling technology to assist plant breeders with the development of improved cultivars for farmers. Yet, despite much investment, there are few examples demonstrating the application of phenomics within a plant breeding program. We review recent progress in field phenomics and highlight the importance of targeting breeders’ needs, rather than perceived technology needs, through developing and enhancing partnerships between phenomics researchers and plant breeders

Caries associated with orthodontic care part 1: aetiology, prevalence and prevention

The prevalence of demineralization, or enamel caries which is the first clinically recognizable stage of caries, has been reported to be as high as 73% during fixed appliance treatment. It is recognized that wearing an orthodontic appliance increases the caries risk of the individual. The orthodontist can effectively reduce demineralization by applying fluoride varnish at each visit. Selecting patients on their oral hygiene clearly is one factor which can prevent patients suffering from demineralization. Excellent home care with optimal use of fluoride will also help prevent the condition. There is little evidence that fluoride-releasing materials used for bonding orthodontic appliances offer protection from demineralization. The first of these two papers describes the aetiology, prevalence and prevention of demineralization associated with orthodontic care. The second paper will describe the treatments available to manage caries focusing on demineralization post orthodontic treatment. CPD/Clinical Relevance: Demineralization is the most common complication of orthodontic care. The clinician should understand how to prevent this

Professionalism and person-centredness: developing a practice based approach to leadership within NHS maternity services in the UK

This paper, based on data taken from in-depth interviews with senior midwives and obstetricians and conducted as part of a critical ethnographic study, argues for a greater appreciation of person-centred, value-led midwifery practice. The paper begins with a discussion of the way midwifery practice is shaped by encoded and embodied knowledge. The paper subsequently focuses on an emergent practice based leadership using an adapted Aristotelian conceptual framework derived from MacIntyre (2007). Professional dissonance is highlighted as a difficulty experienced by repositioned managers who are also expected to be leaders in their field. Using data gathered from in-depth interviews it is contended that establishing person-centred care might be better achieved through the development of practice based leadership (rather than solely by adherence to organisational requirements). This type of leadership could potentially nurture a professional environment that promotes qualities, such as agency, commitment and high levels of competence among midwives. Such leadership is central to UK government priorities and is applicable to a global practice development agenda

Teaching about obesity: Caring, compassion, communication and courage in midwifery education

Teaching innovation can be used to promote the 6Cs in one of the most purportedly stigmatising areas of maternity care: obesity. As rates of maternal obesity continue to rise, getting this area of care right becomes more urgent. Although a great deal has been published about the unsatisfactory and stigmatising impact that staff attitudes can have on those with obesity, there is relatively little on how this problematic area of care should be taught to the next generation of midwives. This article presents a case study of learning about obesity management at the pre-registration, undergraduate level. The case study described is an effort to move away from what is currently largely an academic debate, towards a set of tangible practice recommendations

Tourism destination modelling: building a sustainable planning tool for Australian tourism destinations

The Ningaloo Destination Model is a tourism planning tool for the Ningaloo Coast region of Western Australia that assesses the economic, social and environmental impacts of different planning decisions and events. This report describes the features of the tourism destination model, and analyses its application in the region and to other parts of Australia. Destination modelling integrates a number of research methodologies developed through past STCRC projects (on visitor spending and characteristics, social impacts and economic impacts), secondary data and ecological research. The key to this process is a model development technique that uses scenario planning methodologies to facilitate stakeholder engagement and conceptual modelling techniques to facilitate research integration. The report describes the methodologies used for model development and for data collection, provides two case studies demonstrating outputs, and explores applications of the Ningaloo Destination Model to the region, to other locations and to other sectors. The Ningaloo Destination Model provides estimates of the impacts of plans and events related to tourism in four dimensions: tourism specific; economy; social; and environmental (both resource use and ecological). These outputs are explored through two case studies: a nodal coastal development; and a large resort development. The model can be used for four broad (oftentimes overlapping) categories of assessment: operational planning and decision making for specific organisations and groups (such as local government or agencies that manage land or sea use), regional planning, participatory planning and collaborations and to assist monitoring and evaluation.The Ningaloo Destination Model will be available to the general public (in a limited format) through websites; to agencies through a desktop version; and through integration into a larger model of the region being developed by the CSIRO. Destination modelling is relevant for other tourism destinations and a process for the rapid and cost-effective application of destination modelling is feasible. While much of the data is available, a broader roll-out would require benchmarking of water, electricity and waste data, and developing a wireframe for all tourism destinations. Making destination modelling tools broadly available would significantly broaden the impacts considered in tourism planning and lead to enhancement of desirable effects of tourism development, and early mitigation of negative impacts across Australia. The techniques developed for destination modelling were also found to be applicable to other sectors

Structural evidence for the partially oxidized dipyrromethene and dipyrromethanone forms of the cofactor of porphobilinogen deaminase: structures of the Bacillus megaterium enzyme at near-atomic resolution.

The enzyme porphobilinogen deaminase (PBGD; hydroxymethylbilane synthase; EC 2.5.1.61) catalyses an early step of the tetrapyrrole-biosynthesis pathway in which four molecules of the monopyrrole porphobilinogen are condensed to form a linear tetrapyrrole. The enzyme possesses a dipyrromethane cofactor, which is covalently linked by a thioether bridge to an invariant cysteine residue (Cys241 in the Bacillus megaterium enzyme). The cofactor is extended during the reaction by the sequential addition of the four substrate molecules, which are released as a linear tetrapyrrole product. Expression in Escherichia coli of a His-tagged form of B. megaterium PBGD has permitted the X-ray analysis of the enzyme from this species at high resolution, showing that the cofactor becomes progressively oxidized to the dipyrromethene and dipyrromethanone forms. In previously solved PBGD structures, the oxidized cofactor is in the dipyromethenone form, in which both pyrrole rings are approximately coplanar. In contrast, the oxidized cofactor in the B. megaterium enzyme appears to be in the dipyrromethanone form, in which the C atom at the bridging α-position of the outer pyrrole ring is very clearly in a tetrahedral configuration. It is suggested that the pink colour of the freshly purified protein is owing to the presence of the dipyrromethene form of the cofactor which, in the structure reported here, adopts the same conformation as the fully reduced dipyrromethane form

Effectiveness, Costs and Patient Acceptance of a Conventional and a Biological Treatment Approach for Carious Primary Teeth in Children

BACKGROUND: Over the last years, conventional restorations for the treatment of active carious lesions (CL) in primary teeth have been challenged and a more biological approach has been suggested. This approach involves less invasive techniques that alter the environment of the CL isolating it from the cariogenic biofilm and substrate. AIM: To investigate the cost-effectiveness and patient acceptance of 2 treatment approaches for the treatment of deep CLs in primary teeth in children. METHODS: This was a retrospective/prospective cohort study carried out in 2 UK specialist hospital settings. Data on cost-effectiveness was extracted retrospectively from clinical dental records of 246 patients aged 4-9 years. A prospective study design was used to explore patient acceptance of the 2 treatment approaches. One hundred and ten patients aged 4-9 years and their carers completed 2 questionnaires on treatment acceptance. RESULTS: In total, 836 primary teeth that had received treatment with either approach were included. More than 2 thirds (75.7%) of the restorations in the conventional approach were of non-selective removal to hard dentine followed by pulpotomy (24.3%). In the biological approach, most of the restorations were stainless steel crowns placed with the Hall Technique (95%) followed by selective removal to firm dentine (5%). The majority of the primary teeth remained asymptomatic after a follow-up period of up to 77 months; 95.3% in the conventional and 95.8% in the biological arm. When the treatment costs were analysed, a statistically significant difference was found between the mean costs of the 2 approaches with a mean difference of GBP 45.20 (Pound Sterling; p < 0.001), in favour of the biological approach. The majority of the children and carers were happy with the conventional or biological restorations. CONCLUSION: Although both approaches had similar successful outcomes, the biological approach consisting mainly of Hall Technique was associated with reduced treatment costs. Both approaches were accepted favourably by the children and carers

Live Reality Television: care structures within the production and reception of talent shows

This article focuses on production and reception practices for live reality television, using critical theory and empirical research to question how producers and audiences co-create and limit live experiences. The concept of care structures is used to make visible hidden labour in the creation of mood, in particular audiences as participants in the management of live experiences. In the case of Got to Dance there was a play off between the value and meaning of the live events as a temporary experience captured by ratings and social media, and the more enduring collective-social experience of this reality series over time

Spectral Libraries for SWATH-MS Assays for Drosophila melanogaster and Solanum lycopersicum.

Quantitative proteomics methods have emerged as powerful tools for measuring protein expression changes at the proteome level. Using MS-based approaches, it is now possible to routinely quantify thousands of proteins. However, prefractionation of the samples at the protein or peptide level is usually necessary to go deep into the proteome, increasing both MS analysis time and technical variability. Recently, a new MS acquisition method named SWATH is introduced with the potential to provide good coverage of the proteome as well as a good measurement precision without prior sample fractionation. In contrast to shotgun-based MS however, a library containing experimental acquired spectra is necessary for the bioinformatics analysis of SWATH data. In this study, spectral libraries for two widely used models are built to study crop ripening or animal embryogenesis, Solanum lycopersicum (tomato) and Drosophila melanogaster, respectively. The spectral libraries comprise fragments for 5197 and 6040 proteins for S. lycopersicum and D. melanogaster, respectively, and allow reproducible quantification for thousands of peptides per MS analysis. The spectral libraries and all MS data are available in the MassIVE repository with the dataset identifiers MSV000081074 and MSV000081075 and the PRIDE repository with the dataset identifiers PXD006493 and PXD006495