¹H, ¹⁵N, ¹³C backbone resonance assignments of human phosphoglycerate kinase in a transition state analogue complex with ADP, 3-phosphoglycerate and magnesium trifluoride

Human phosphoglycerate kinase (PGK) is an energy generating glycolytic enzyme that catalyses the transfer of a phosphoryl group from 1,3-bisphosphoglycerate (BPG) to ADP producing 3-phosphoglycerate (3PG) and ATP. PGK is composed of two α/β Rossmann-fold domains linked by a central α-helix and the active site is located in the cleft formed between the N-domain which binds BPG or 3PG, and the C-domain which binds the nucleotides ADP or ATP. Domain closure is required to bring the two substrates into close proximity for phosphoryl transfer to occur, however previous structural studies involving a range of native substrates and substrate analogues only yielded open or partly closed PGK complexes. X-ray crystallography using magnesium trifluoride (MgF3(-)) as a isoelectronic and near-isosteric mimic of the transferring phosphoryl group (PO3(-)), together with 3PG and ADP has been successful in trapping human PGK in a fully closed transition state analogue (TSA) complex. In this work we report the (1)H, (15)N and (13)C backbone resonance assignments of human PGK in the solution conformation of the fully closed PGK:3PG:MgF3:ADP TSA complex. Assignments were obtained by heteronuclear multidimensional NMR spectroscopy. In total, 97% of all backbone resonances were assigned in the complex, with 385 out of a possible 399 residues assigned in the (1)H-(15)N TROSY spectrum. Prediction of solution secondary structure from a chemical shift analysis using the TALOS-N webserver is in good agreement with the published X-ray crystal structure of this complex

The impact of gastrointestinal parasitism on the behaviour and welfare of weaned housed lambs

Gastrointestinal (GI) parasitism is a health and production concern in sheep, yet its impact on animal welfare remains unclear. The impact of subclinical infections is especially ambiguous as GI parasitism often remains undiagnosed until clinical signs such as diarrhoea are evident. This study applied quantitative and qualitative methods to examine the effects of subclinical Teladorsagia circumcincta infection on the behaviour and welfare of 96 Suffolk-cross lambs (24 pens of 4 lambs) weaned at 10 weeks old. The hypothesis that parasitism causes negative affective states was tested. Lambs were divided into three groups at the pen level: ad-lib fed control (AC), restricted-fed control (RC), and ad-lib fed parasitised (AP). Parasitised lambs (AP) were dosed three times weekly with 7000 third stage T. circumcincta larvae (L 3) from 16 weeks of age. Lambs in the RC group were pair fed to match AP feed intake to separate the effects of infection-induced anorexia from the potential direct impacts of infection. From 7 days pre-infection to 23 days post-infection, scan and behaviour samples were taken from video recordings to quantitatively monitor behaviour, and animal-based measures such as faecal soiling score (FSS) were recorded as welfare indicators. Lying, standing, eating, play and social behaviour were monitored. Qualitative behaviour assessment (QBA) was conducted weekly using the AWIN (2015) protocol to gain insight into the lambs’ affective states over the onset of infection. Parasitised lambs were more likely to stand inactive than AC lambs as the infection progressed (P=0.006). They were also less likely to display eating behaviour in the third daily scan sample than RC lambs (P&lt;0.001). Principal Component Analysis of the QBA data revealed that the first dimension (PC1) described arousal levels, the second (PC2) described the valence of the animals’ affective states, and the third (PC3) described fearfulness and aggression levels. Parasitised lambs (est=10.64,SE=0.33) scored higher than RC lambs (est=9.42, SE=0.33) on PC3, the fearfulness dimension (P=0.030). There were no differences between fearfulness scores of AC and AP lambs or RC lambs and treatment group had no significant impact on the distribution of scores on PC1 or PC2. These findings demonstrate that subclinical GI parasitism negatively impacts lamb welfare not only in the health domain but in the behaviour and mental domains as well. This has implications for welfare assessments and early disease detection in lambs. Future research could explore remote monitoring of the indicators of parasitism identified in this study.</p

The impact of gastrointestinal parasitism on the behaviour and welfare of weaned housed lambs

Gastrointestinal (GI) parasitism is a health and production concern in sheep, yet its impact on animal welfare remains unclear. The impact of subclinical infections is especially ambiguous as GI parasitism often remains undiagnosed until clinical signs such as diarrhoea are evident. This study applied quantitative and qualitative methods to examine the effects of subclinical Teladorsagia circumcincta infection on the behaviour and welfare of 96 Suffolk-cross lambs (24 pens of 4 lambs) weaned at 10 weeks old. The hypothesis that parasitism causes negative affective states was tested. Lambs were divided into three groups at the pen level: ad-lib fed control (AC), restricted-fed control (RC), and ad-lib fed parasitised (AP). Parasitised lambs (AP) were dosed three times weekly with 7000 third stage T. circumcincta larvae (L 3) from 16 weeks of age. Lambs in the RC group were pair fed to match AP feed intake to separate the effects of infection-induced anorexia from the potential direct impacts of infection. From 7 days pre-infection to 23 days post-infection, scan and behaviour samples were taken from video recordings to quantitatively monitor behaviour, and animal-based measures such as faecal soiling score (FSS) were recorded as welfare indicators. Lying, standing, eating, play and social behaviour were monitored. Qualitative behaviour assessment (QBA) was conducted weekly using the AWIN (2015) protocol to gain insight into the lambs’ affective states over the onset of infection. Parasitised lambs were more likely to stand inactive than AC lambs as the infection progressed (P=0.006). They were also less likely to display eating behaviour in the third daily scan sample than RC lambs (P&lt;0.001). Principal Component Analysis of the QBA data revealed that the first dimension (PC1) described arousal levels, the second (PC2) described the valence of the animals’ affective states, and the third (PC3) described fearfulness and aggression levels. Parasitised lambs (est=10.64,SE=0.33) scored higher than RC lambs (est=9.42, SE=0.33) on PC3, the fearfulness dimension (P=0.030). There were no differences between fearfulness scores of AC and AP lambs or RC lambs and treatment group had no significant impact on the distribution of scores on PC1 or PC2. These findings demonstrate that subclinical GI parasitism negatively impacts lamb welfare not only in the health domain but in the behaviour and mental domains as well. This has implications for welfare assessments and early disease detection in lambs. Future research could explore remote monitoring of the indicators of parasitism identified in this study.</p

Why the energy landscape of barnase is hierarchical

We have used NMR and computational methods to characterize the dynamics of the ribonuclease barnase over a wide range of timescales in free and inhibitor-bound states. Using temperature- and denaturant-dependent measurements of chemical shift, we show that barnase undergoes frequent and highly populated hinge bending. Using relaxation dispersion, we characterize a slower and less populated motion with a rate of 750 ± 200 s-¹, involving residues around the lip of the active site, which occurs in both free and bound states and therefore suggests conformational selection. Normal mode calculations characterize correlated hinge bending motions on a very rapid timescale. These three measurements are combined with previous measurements and molecular dynamics calculations on barnase to characterize its dynamic landscape on timescales from picoseconds to milliseconds and length scales from 0.1 to 2.5 nm. We show that barnase has two different large-scale fluctuations: one on a timescale of 10-⁹-10-⁶ s that has no free energy barrier and is a hinge bending that is determined by the architecture of the protein; and one on a timescale of milliseconds (i.e., 750 s-¹) that has a significant free energy barrier and starts from a partially hinge-bent conformation. These two motions can be described as hierarchical, in that the more highly populated faster motion provides a platform for the slower (less probable) motion. The implications are discussed. The use of temperature and denaturant is suggested as a simple and general way to characterize motions on the intermediate ns-μs timescale

A timeband framework for modelling real-time systems

Complex real-time systems must integrate physical processes with digital control, human operation and organisational structures. New scientific foundations are required for specifying, designing and implementing these systems. One key challenge is to cope with the wide range of time scales and dynamics inherent in such systems. To exploit the unique properties of time, with the aim of producing more dependable computer-based systems, it is desirable to explicitly identify distinct time bands in which the system is situated. Such a framework enables the temporal properties and associated dynamic behaviour of existing systems to be described and the requirements for new or modified systems to be specified. A system model based on a finite set of distinct time bands is motivated and developed in this paper

Physical activity as a treatment for depression: the TREAD randomised trial protocol

Published version. Copyright © 2010 BioMed CentralBackground: Depression is one of the most common reasons for consulting a General Practitioner (GP) within the UK. Whilst antidepressants have been shown to be clinically effective, many patients and healthcare professionals would like to access other forms of treatment as an alternative or adjunct to drug therapy for depression. A recent systematic review presented some evidence that physical activity could offer one such option, although further investigation is needed to test its effectiveness within the context of the National Health Service. The aim of this paper is to describe the protocol for a randomised, controlled trial (RCT) designed to evaluate an intervention developed to increase physical activity as a treatment for depression within primary care. Methods/design: The TREAD study is a pragmatic, multi-centre, two-arm RCT which targets patients presenting with a new episode of depression. Patients were approached if they were aged 18-69, had recently consulted their GP for depression and, where appropriate, had been taking antidepressants for less than one month. Only those patients with a confirmed diagnosis of a depressive episode as assessed by the Clinical Interview Schedule-Revised (CIS-R), a Beck Depression Inventory (BDI) score of at least 14 and informed written consent were included in the study. Eligible patients were individually randomised to one of two treatment groups; usual GP care or usual GP care plus facilitated physical activity. The primary outcome of the trial is clinical symptoms of depression assessed using the BDI four months after randomisation. A number of secondary outcomes are also measured at the 4-, 8- and 12-month follow-up points including quality of life, attitude to and involvement in physical activity and antidepressant use/adherence. Outcomes will be analysed on an intention-to-treat (ITT) basis and will use linear and logistic regression models to compare treatments. Discussion: The results of the trial will provide information about the effectiveness of physical activity as a treatment for depression. Given the current prevalence of depression and its associated economic burden, it is hoped that TREAD will provide a timely contribution to the evidence on treatment options for patients, clinicians and policy-makers

Allomorphy as a mechanism of post-translational control of enzyme activity

Enzyme regulation is vital for metabolic adaptability in living systems. Fine control of enzyme activity is often delivered through post-translational mechanisms, such as allostery or allokairy. β-phosphoglucomutase (βPGM) from Lactococcus lactis is a phosphoryl transfer enzyme required for complete catabolism of trehalose and maltose, through the isomerisation of β-glucose 1-phosphate to glucose 6-phosphate via β-glucose 1,6-bisphosphate. Surprisingly for a gatekeeper of glycolysis, no fine control mechanism of βPGM has yet been reported. Herein, we describe allomorphy, a post-translational control mechanism of enzyme activity. In βPGM, isomerisation of the K145-P146 peptide bond results in the population of two conformers that have different activities owing to repositioning of the K145 sidechain. In vivo phosphorylating agents, such as fructose 1,6-bisphosphate, generate phosphorylated forms of both conformers, leading to a lag phase in activity until the more active phosphorylated conformer dominates. In contrast, the reaction intermediate β-glucose 1,6-bisphosphate, whose concentration depends on the β-glucose 1-phosphate concentration, couples the conformational switch and the phosphorylation step, resulting in the rapid generation of the more active phosphorylated conformer. In enabling different behaviours for different allomorphic activators, allomorphy allows an organism to maximise its responsiveness to environmental changes while minimising the diversion of valuable metabolites

Consumer perceptions of co-branding alliances: Organizational dissimilarity signals and brand fit

This study explores how consumers evaluate co-branding alliances between dissimilar partner firms. Customers are well aware that different firms are behind a co-branded product and observe the partner firms’ characteristics. Drawing on signaling theory, we assert that consumers use organizational characteristics as signals in their assessment of brand fit and for their purchasing decisions. Some organizational signals are beyond the control of the co-branding partners or at least they cannot alter them on short notice. We use a quasi-experimental design and test how co-branding partner dissimilarity affects brand fit perception. The results show that co-branding partner dissimilarity in terms of firm size, industry scope, and country-of-origin image negatively affects brand fit perception. Firm age dissimilarity does not exert significant influence. Because brand fit generally fosters a benevolent consumer attitude towards a co-branding alliance, the findings suggest that high partner dissimilarity may reduce overall co-branding alliance performance

The global impact of offshore wind farms on ecosystem services

Understanding the global impact of offshore wind farms (OWF) on biodiversity and ecosystem services (ES) is crucial in developing sustainable energy transition pathways. This study takes a holistic approach, coupling a semi-systematic review with a novel analytical methodology, to consider the consequences of construction & operation of OWF deployment on biodiversity and ES. 314 pieces of evidence taken from 132 peer-reviewed studies provide the basis to determine the ecological and ES impacts. The process showed that construction impacts were predominantly negative across the ecological subject groups (52%), compared with positive im�pacts (8%) with several species of fish (e.g. brill, cod, dab, plaice) and some species of birds (e.g. common guillemot, northern fulmar, redhead) showing strongly negative trends. Operational phase impacts were more variable and could be either negative (32%) or positive (34%) depending on site specific conditions. More detailed investigations into fish, shellfish, humans and air-surface studies recorded a net positive effect of wind farm operations on these subject groups. Translation into ES outcomes identified that 14 ES are impacted by the construction and operation of OWF. The most substantially enhanced ES included effects on commercial fisheries and experiential recreation. Social acceptance toward new and hypothetical OWF was also strongly positive, irrespective of country location. Negative effects on ES, including existence values for culturally important groups, e.g., marine mammals and birds and the spread of non-native species, are potentially of most signifi�cance. Overall, this study finds more than 86% of possible offshore wind farm impacts on ES are still unknown. There was also a paucity of studies on the decommissioning of OWF and the impacts of deeper-water floating structures, with a bias in studies toward northern hemisphere and developed countries

Adaptive Introgression across Species Boundaries in Heliconius Butterflies

It is widely documented that hybridisation occurs between many closely related species, but the importance of introgression in adaptive evolution remains unclear, especially in animals. Here, we have examined the role of introgressive hybridisation in transferring adaptations between mimetic Heliconius butterflies, taking advantage of the recent identification of a gene regulating red wing patterns in this genus. By sequencing regions both linked and unlinked to the red colour locus, we found a region that displays an almost perfect genotype by phenotype association across four species, H. melpomene, H. cydno, H. timareta, and H. heurippa. This particular segment is located 70 kb downstream of the red colour specification gene optix, and coalescent analysis indicates repeated introgression of adaptive alleles from H. melpomene into the H. cydno species clade. Our analytical methods complement recent genome scale data for the same region and suggest adaptive introgression has a crucial role in generating adaptive wing colour diversity in this group of butterflies