Radiation damage to nucleoprotein complexes in macromolecular crystallography

Significant progress has been made in macromolecular crystallography over recent years in both the understanding and mitigation of X-ray induced radiation damage when collecting diffraction data from crystalline proteins. In contrast, despite the large field that is productively engaged in the study of radiation chemistry of nucleic acids, particularly of DNA, there are currently very few X-ray crystallographic studies on radiation damage mechanisms in nucleic acids. Quantitative comparison of damage to protein and DNA crystals separately is challenging, but many of the issues are circumvented by studying pre-formed biological nucleoprotein complexes where direct comparison of each component can be made under the same controlled conditions. Here a model protein-DNA complex C.Esp1396I is employed to investigate specific damage mechanisms for protein and DNA in a biologically relevant complex over a large dose range (2.07-44.63 MGy). In order to allow a quantitative analysis of radiation damage sites from a complex series of macromolecular diffraction data, a computational method has been developed that is generally applicable to the field. Typical specific damage was observed for both the protein on particular amino acids and for the DNA on, for example, the cleavage of base-sugar N1-C and sugar-phosphate C-O bonds. Strikingly the DNA component was determined to be far more resistant to specific damage than the protein for the investigated dose range. At low doses the protein was observed to be susceptible to radiation damage while the DNA was far more resistant, damage only being observed at significantly higher doses

Big Hole (41TV2161): Two Stratigraphically Isolated Middle Holocene Components in Travis County, Texas Volume I

During April and May 2006, an archeological team from the Cultural Resources Section of the Planning, Permitting and Licensing Practice of TRC Environmental Corporation’s (TRC) Austin office conducted geoarcheological documentation and data recovery excavations at prehistoric site 41TV2161 (CSJ: 0440-06-006). Investigations were restricted to a 70 centimeter (cm) thick target zone between ca. 220 and 290 cm below surface (bs) on the western side of site 41TV2161 – the Big Hole site in eastern Travis County, Texas. This cultural investigation was necessary under the requirements of Section 106 of the National Historic Preservation Act (NHPA), the implementing regulations of 36CRF Part 800 and the Antiquities Code of Texas (Texas Natural Resource Code, Title 9, Chapter 191 as amended) to recover a sample of the significant cultural materials prior to destruction by planned construction of State Highway 130 (SH 130). The latter by a private construction firm – Lone Star Infrastructure. This necessary data recovery was for Texas Department of Transportation (TxDOT), Environmental (ENV) Affairs Division under a Scientific Services Contract No. 577XXSA003 (Work Authorization No. 57701SA003). Over the years since the original award, multiple work authorizations between TxDOT and TRC were implemented and completed towards specific aspects of the analyses and reporting. The final analyses and report were conducted under contract 57-3XXSA004 (Work Authorization 57-311SA004). All work was under Texas Antiquities Committee Permit No. 4064 issued by the Texas Historical Commission (THC) to J. Michael Quigg. Initially, an archeological crew from Hicks & Company encountered site 41TV2161 during an intensive cultural resource inventory conducted south of Pearce Lane along the planned construction zone of SH 130 in the fall of 2005. Following the initial site discovery, archeologists expanded their investigations to the west across the SH 130 right-of-way, and completed excavation of 10 backhoe trenches, 13 shovel tests, and 11 test units at site 41TV2161. The investigations encountered at least seven buried cultural features and 1,034 artifacts, some in relatively good context. The survey and testing report to TxDOT presented their findings and recommendations (Campbell et al. 2006). The ENV Affairs Division of TxDOT and the THC reviewed the initial findings and recommendations, and determined site 41TV2161 was eligible for listing on the National Register of Historic Places and as State Antiquities Landmark as the proposed roadway development was to directly impact this important site and further excavations were required. Subsequently, TRC archeologists led by Paul Matchen (Project Archeologist) and J. Michael Quigg (Principal Investigator) initiated data recovery excavations through the mechanical-removal of between 220 and 250 cm of sediment from a 30-by-40 meter (m) block area (roughly 3,000 m3). This was conducted to allow hand-excavations to start just above the deeply buried, roughly 70 cm thick targeted zone of cultural material. Mechanical stripping by Lone Star Infrastructure staff created a large hole with an irregular bottom that varied between 220 and 260 cmbs. To locate specific areas to initiate hand-excavations within the mechanically stripped area, a geophysical survey that employed ground penetrating radar (GPR) was conducted by Tiffany Osburn then with Geo-Marine in Plano, Texas. Over a dozen electronic anomalies were detected through the GPR investigation. Following processing, data filtering, and assessment, Osburn identified and ranked the anomalies for investigation. The highest ranked anomalies (1 through 8) were thought to have the greatest potential to represent cultural features. Anomalies 1 through 6 were selected and targeted through hand-excavations of 1-by-1 m units that formed continuous excavation blocks of various sizes. Blocks were designated A, B, C, D, E, and F. The type, nature, quantity, and context of encountered cultural materials in each block led the direction and expansion of each excavation block as needed. In total, TRC archeologists hand-excavated 38.5 m3 (150 m2) from a vertically narrow target zone within this deep, multicomponent and stratified prehistoric site. Hand-excavation in the two largest Blocks, B and D (51 m2 and 62 m2 respectively), revealed two vertically separate cultural components between roughly 220 and 290 cmbs. The younger component was restricted to Block B and yielded a Bell/Andice point and point base, plus a complete Big Sandy point. These points were associated with at least eight small burned rock features, one cluster of ground stone tools, limited quantities of lithic debitage, few formal chipped and ground stone tools, and a rare vertebrate faunal assemblage. Roughly 20 to 25 cm below the Bell/Andice component in Block B and across Block D was a component identified by a single corner-notched Martindale dart point. This point was associated with a scattered burned rocks, three charcoal stained hearth features, scattered animal, bird, and fish bones, mussel shells, and less than a dozen formal chipped and ground stone tools. Both identified components contained cultural materials in good stratigraphic context with high spatial integrity. Significant, both were radiocarbon dated by multiple charcoal samples to a narrow 200-year period between 5250 and 5450 B.P. during the middle Holocene. With exception of the well-preserved faunal assemblages, perishable materials were poorly preserved in the moist silty clay loam. Charcoal lacked structure and was reduced to dark stains. Microfossils (e.g., phytoliths and starch gains) were present, although in very limited numbers and deteriorated conditions. The four much smaller Blocks (A, C, E, and F) yielded various quantities of cultural material and features, but these blocks also lacked sufficient charcoal dates and diagnostic artifacts Those artifacts and samples were left unassigned and analyzed separately from the Bell/Andice and Martindale components. The two well-defined components in Blocks B and D are the focus of this technical report. The components provide very significant data towards understanding rare and poorly understood hunter-gatherer populations during late stages of the Altithermal climate period. This final report builds upon the interim report submitted to TxDOT (Quigg et al. 2007) that briefly described the methods, excavations, preliminary findings, initial results from six feasibility studies, and proposed an initial research design for data analyses. Context and integrity of the cultural materials in the two identified components was excellent. This rare circumstance combined with detailed artifact analyses, solid documentation of their ages through multiple radiocarbon dates, and multidisciplinary approach to analyses, allowed significant insights and contributions concerning the two populations involved. Results provide a greater understanding of human behaviors during a rarely identified time in Texas Prehistory. The cultural materials and various collected samples were temporarily curated at TRC’s Austin laboratory. Following completion of analyses and acceptance of this final report, the artifacts, paper records, photographs, and electronic database were permanently curated at the Center for Archaeological Studies (CAS) at Texas State University in San Marcos

Life on a slippery slope: perceptions of health in adults with cystic fibrosis

This paper focuses on how adults with cystic fibrosis (CF) attending a specialist CF centre in the UK perceive their health. In common with many other genetic diseases, CF is traditionally conceptualised as a fatal childhood disease, yet the average survival age for those with CF has been steadily rising over the past half century. Thus it is now predicted that those born in 1990 will live on average for 40 years. To date, however, most sociological work has focused on children or adolescents affected by CF rather than on adults between the ages of 18 and 40, the focus of the study reported here. The paper shows that these adults' varying perceptions of health are related to the effects of CF, its treatment, and the context in which adults are placed. Four concepts of health are identified (health as 'normal', controllable, distressing and a release) along with certain styles, ways of coping and related strategies. Through these analytic distinctions the paper aims to make a contribution to the sociological understanding of lay concepts of health in adults with childhood or genetic disease

RNA protects a nucleoprotein complex against radiation damage

Radiation damage during macromolecular X-ray crystallographic data collection is still the main impediment for many macromolecular structure determinations. Even when an eventual model results from the crystallographic pipeline, the manifestations of radiation-induced structural and conformation changes, the so-called specific damage, within crystalline macromolecules can lead to false interpretations of biological mechanisms. Although this has been well characterized within protein crystals, far less is known about specific damage effects within the larger class of nucleoprotein complexes. Here, a methodology has been developed whereby per-atom density changes could be quantified with increasing dose over a wide (1.3-25.0 MGy) range and at higher resolution (1.98 Å) than the previous systematic specific damage study on a protein-DNA complex. Specific damage manifestations were determined within the large trp RNA-binding attenuation protein (TRAP) bound to a single-stranded RNA that forms a belt around the protein. Over a large dose range, the RNA was found to be far less susceptible to radiation-induced chemical changes than the protein. The availability of two TRAP molecules in the asymmetric unit, of which only one contained bound RNA, allowed a controlled investigation into the exact role of RNA binding in protein specific damage susceptibility. The 11-fold symmetry within each TRAP ring permitted statistically significant analysis of the Glu and Asp damage patterns, with RNA binding unexpectedly being observed to protect these otherwise highly sensitive residues within the 11 RNA-binding pockets distributed around the outside of the protein molecule. Additionally, the method enabled a quantification of the reduction in radiation-induced Lys and Phe disordering upon RNA binding directly from the electron density

Development of tools to automate quantitative analysis of radiation damage in SAXS experiments

Biological small-angle X-ray scattering (SAXS) is an increasingly popular technique used to obtain nanoscale structural information on macromolecules in solution. However, radiation damage to the samples limits the amount of useful data that can be collected from a single sample. In contrast to the extensive analytical resources available for macromolecular crystallography (MX), there are relatively few tools to quantitate radiation damage for SAXS, some of which require a significant level of manual characterization, with the potential of leading to conflicting results from different studies. Here, computational tools have been developed to automate and standardize radiation damage analysis for SAXS data. RADDOSE-3D, a dose calculation software utility originally written for MX experiments, has been extended to account for the cylindrical geometry of the capillary tube, the liquid composition of the sample and the attenuation of the beam by the capillary material to allow doses to be calculated for many SAXS experiments. Furthermore, a library has been written to visualize and explore the pairwise similarity of frames. The calculated dose for the frame at which three subsequent frames are determined to be dissimilar is defined as the radiation damage onset threshold (RDOT). Analysis of RDOTs has been used to compare the efficacy of radioprotectant compounds to extend the useful lifetime of SAXS samples. Comparison of the RDOTs shows that, for radioprotectant compounds at 5 and 10 mM concentration, glycerol is the most effective compound. However, at 1 and 2 mM concentrations, di­thio­threitol (DTT) appears to be most effective. Our newly developed visualization library contains methods that highlight the unusual radiation damage results given by SAXS data collected using higher concentrations of DTT: these observations should pave the way to the development of more sophisticated frame merging strategies

'Experts', 'partners' and 'fools': exploring agency in HIV treatment seeking among African migrants in London

In an attempt to promote patient agency and foster more egalitarian relationships between patients and doctors, discourse concerning health and wellbeing in the UK has increasingly centred around the notion of informed and 'expert' patients who are able to effectively input into the direction and management of their own health care and treatment. While the relationship between a patient and their doctor can play a vital role in influencing the treatment decisions and health-related outcomes of people living with long term illness, little is known about the ways in which people living with HIV actually perceive their relationship with their doctors, nor the implications this may have for the types of treatment they may seek to use and the related information that they share. Drawing on 11 focus group discussions and 20 repeat interviews undertaken in 2008-2009 with HIV-positive adult migrants from Zambia, Zimbabwe and South Africa living in the UK, this paper argues that patient-doctor relationships can be heavily influenced by the perceived legitimacy of different forms of medical knowledge and treatments and by culturally influenced ideas regarding health, wellbeing and agency. Despite a desire amongst some migrants to use 'traditional' medicines from southern Africa as well as other non-biomedical treatments and therapies, the research found that the perceived lack of legitimacy associated with these treatments in the UK rendered their use a largely clandestine activity. At the same time, many patients made clear distinctions concerning issues affecting their immediate health and factors influencing their more general wellbeing, which in turn, impacted upon the information that they chose to share with, or conceal from, their doctors. Such findings challenge assumptions underpinning policy promoting patient agency and have significant and, in cases, potentially adverse implications for the safety and effective administration and management of HIV treatments in African migrant populations and possibly more generally. (C) 2009 Elsevier Ltd. All rights reserved

Advanced Lithium Ion Venus Explorer (ALIVE)

The COncurrent Multidisciplinary Preliminary Assessment of Space Systems (COMPASS) Team partnered with the Applied Research Laboratory to perform a NASA Innovative Advanced Concepts (NIAC) Program study to evaluate chemical based power systems for keeping a Venus lander alive (power and cooling) and functional for a period of days. The mission class targeted was either a Discovery ($500M) or New Frontiers ($750M to $780M) class mission. Historic Soviet Venus landers have only lasted on the order of 2 hours in the extreme Venus environment:temperatures of 460 degrees Centigrade and pressures of 93 bar. Longer duration missions have been studied using plutonium powered systems to operate and cool landers for up to a year. However, the plutonium load is very large. This NIAC study sought to still provide power and cooling but without the plutonium. Batteries are far too heavy but a system which uses the atmosphere (primarily carbon dioxide) and on on-board fuel to power a power generation and cooling system was sought. The resuling design was the Advanced Long-Life Lander Investigating the Venus Environment (ALIVE) Spacecraft (S/C) which burns lithium (Li) with the CO2 atmosphere to heat a Duplex Stirling to power and cool the lander for a 5-day duration (until the Li is exhausted). While it does not last years a chemical powered system surviving days eliminates the cost associated with utilizing a flyby relay S/C and allows a continuous low data rate direct to earth (DTE) link in this instance from the Ovda Regio of Venus. The five-day collection time provided by the chemical power systems also enables science personnel on earth to interact and retarget science - something not possible with an approximately 2-hour spacecraft lifetime. It also allows for contingency operations directed by the ground (reduced risk). The science package was based on that envisioned by the Venus Intrepid Tessera Lander (VITaL) Decadal Survey Study. The Li Burner within the long duration power system creates approximately 14000 W of heat. This 1300 degree Centigrade heat using Li in the bottom "ballast" tank is melted to liquid by the Venus temperature, drawn into a furnace by a wick and burned with atmospheric CO2. The Li carbonate exhaust is liquid at 1300 degrees Centigrade and being denser than Li drains into the the Li tank and solidifies. Since the exhaust product is a dense liquid no "chimney" is required which conserves the heat for the stirling power convertor. The Duplex Stirling provides about 300 W of power and removes about 300 W of heat from the avionics and heat that leaks into the 1-bar-insulated payload pressure vessel kept at 25 degrees Centigrade. The Na K radiator is run to the top of the drag flap.The ALIVE vehicle is carried to Venus via an Atlas 411 launch vehicle (LV) with a C3 of 7 km2/s2. An Aeroshell, derived from the Genesis mission, enables a direct entry into the atmosphere of Venus (-10 degrees Centigrade, 40 g max) and 6 m/s for landing (44 g) using a drag ring. For surface science and communication, a 100 WRF (WebEx Recording Format), X-Band 0.6-meter pointable DTE (Direct-to-Earth) antenna provides 2 kbps (kilobits per second) to DSN (Deep-Space Network) 34-meter antenna clusters.Table 1.1 summarizes the top-level details of each subsystem that was incorporated into the design. Cost estimates of the ALIVE mission show it at approximately$760M which puts it into the New Frontiers class.The ALIVE landed duration is only limited by the amount of Li which can be carried by the lander. Further studies are needed to investigate how additional mass can be carried, perhaps by a larger launcher and larger aeroshell

Molecular mechanisms of toxicity of silver nanoparticles in zebrafish embryos.

addresses: Biosciences, College of Life and Environmental Sciences, Geoffrey Pope Building, University of Exeter, Stocker Road, Exeter, EX4 4QD, UK. [email protected]: Journal Article; Research Support, Non-U.S. Gov'tThis is an open access article that is freely available in ORE or from the publisher's web site. http://pubs.acs.org/doi/abs/10.1021/es401758d. Please cite the published version© 2013 American Chemical SocietySupporting Information: Further details on the methodology and results for the characterization of the silver particles used for the exposures, mortality curves, sequencing analysis, and a number of supporting figures and tables. This material is available free of charge via the Internet at http://pubs.acs.org.Silver nanoparticles cause toxicity in exposed organisms and are an environmental health concern. The mechanisms of silver nanoparticle toxicity, however, remain unclear. We examined the effects of exposure to silver in nano-, bulk-, and ionic forms on zebrafish embryos (Danio rerio) using a Next Generation Sequencing approach in an Illumina platform (High-Throughput SuperSAGE). Significant alterations in gene expression were found for all treatments and many of the gene pathways affected, most notably those associated with oxidative phosphorylation and protein synthesis, overlapped strongly between the three treatments indicating similar mechanisms of toxicity for the three forms of silver studied. Changes in oxidative phosphorylation indicated a down-regulation of this pathway at 24 h of exposure, but with a recovery at 48 h. This finding was consistent with a dose-dependent decrease in oxygen consumption at 24 h, but not at 48 h, following exposure to silver ions. Overall, our data provide support for the hypothesis that the toxicity caused by silver nanoparticles is principally associated with bioavailable silver ions in exposed zebrafish embryos. These findings are important in the evaluation of the risk that silver particles may pose to exposed vertebrate organisms.Natural Environment Research Council (NERC)NERC Biomolecular Analysis FacilityUK Environment AgencySystems Biology Seed fund, University of Exete

Social meanings and understandings in patient-nurse interaction in the community practice setting: a grounded theory study

Provisional: Background: The patient-nurse relationship is a traditional concern of healthcare research. However, patient-nurse interaction is under examined from a social perspective. Current research focuses mostly on specific contexts of care delivery and experience related to medical condition or illness, or to nurses' speciality. Consequentially, this paper is about the social meanings and understandings at play within situated patient-nurse interaction in the community practice setting in a transforming healthcare service. Methods: Grounded theory methodology was used and the research process was characterised by principles of theoretical sensitivity and constant comparative analysis. The field of study was four health centres in the community. The participants were patients and nurses representative of those attending or working in the health centres and meeting there by scheduled appointment. Data collection methods were observations, informal interviews and semi-structured interviews. Results: Key properties of 'Being a good patient, being a good nurse', 'Institutional experiences' and 'Expectations about healthcare' were associated with the construction of a category entitled 'Experience'. Those key properties captured that in an evolving healthcare environment individuals continually re-constructed their reality of being a patient or nurse as they endeavoured to perform appropriately; articulation of past and present healthcare experiences was important in that process. Modus operandi in role as patient was influenced by past experiences in healthcare and by those in non-healthcare institutions in terms of engagement and involvement (or not) in interaction. Patients' expectations about interaction in healthcare included some uncertainly as they strived to make sense of the changing roles and expertise of nurses and, differentiating between the roles and expertise of nurses and doctors. Conclusions: The importance of social meanings and understandings in patient-nurse interaction is not fully apparent to nurses, but important in the patient experience. Seeking understanding from a social perspective makes a contribution to enhancing knowledge about patient-nurse interaction with subsequent impact on practice, in particular the development of the patient-nurse relationship. The implications are that the meanings and understandings patients and nurses generate from experiences beyond and within their situated interaction are pivotal to the development of their relationship in the transforming community healthcare environment

A randomised controlled trial, cost-effectiveness and process evaluation of the implementation of self-management for chronic gastrointestinal disorders in primary care, and linked projects on identification and risk assessment

Background: Chronic gastrointestinal disorders are major burdens in primary care. Although there is some evidence that enhancing self-management can improve outcomes, it is not known if such models of care can be implemented at scale in routine NHS settings and whether or not it is possible to develop effective risk assessment procedures to identify patients who are likely to become chronically ill.Objectives: What is the clinical effectiveness and cost-effectiveness of an intervention to enhance self-management support for patients with chronic conditions when translated from research settings into routine care? What are the barriers and facilitators that affect the implementation of an intervention to enhance self-management support among patients, clinicians and organisations? Is it possible to develop methods to identify patients at risk of long-term problems with functional gastrointestinal disorders in primary care? Data sources included professional and patient interviews, patient self-report measures and data on service utilisation.Design: A pragmatic, two-arm, practice-level cluster Phase IV randomised controlled trial evaluating outcomes and costs associated with the intervention, with associated process evaluation using interviews and other methods. Four studies around identification and risk assessment: (1) a general practitioner (GP) database study to describe how clinicians in primary care record consultations with patients who experience functional lower gastrointestinal symptoms; (2) a validation of a risk assessment tool; (3) a qualitative study to explore GPs’ views and experiences; and (4) a second GP database study to investigate patient profiles in irritable bowel syndrome, inflammatory bowel disease and abdominal pain.Setting: Salford, UK. Participants: People with long-term conditions and professionals in primary care. Interventions: A practice-level intervention to train practitioners to assess patient self-management capabilities and involve them in a choice of self-management options.Main outcome measures: Patient self-management, care experience and quality of life, health-care utilisation and costs.Results: No statistically significant differences were found between patients attending the trained practices and those attending control practices on any of the primary or secondary outcomes. The intervention had little impact on either costs or effects within the time period of the trial. In the practices, self-management tools failed to be normalised in routine care. Full assessment of the predictive tool was not possible because of variable case definitions used in practices. There was a lack of perceived clinical benefit among GPs.Limitations: The intervention was not implemented fully in practice. Assessment of the risk assessment tool faced barriers in terms of the quality of codting in GP databases and poor recruitment of patients.Conclusions: The Whole system Informing Self-management Engagement self-management (WISE) model did not add value to existing care for any of the long-term conditions studied.Future work: The active components required for effective self-management support need further study. The results highlight the challenge of delivering improvements to quality of care for long-term conditions. There is a need to develop interventions that are feasible to deliver at scale, yet demonstrably clinically effective and cost-effective. This may have implications for the piloting of interventions and linking implementation more clearly to local commissioning strategies.<br/