Substrate Engagement and Catalytic Mechanisms of N-Acetylglucosaminyltransferase v

α-Mannoside β-1,6-N-acetylglucosaminyltransferase V (MGAT5) is a mammalian glycosyltransferase involved in complex N-glycan formation, which strongly drives cancer when overexpressed. Despite intense interest, the catalytic mechanism of MGAT5 is not known in detail, precluding therapeutic exploitation. We solved structures of MGAT5 complexed to glycosyl donor and acceptor ligands, revealing an unforeseen role for donor-induced loop rearrangements in controlling acceptor substrate engagement. QM/MM metadynamics simulations of MGAT5 catalysis highlight the key assisting role of Glu297 and reveal considerable conformational distortions imposed upon the glycosyl donor during transfer. Detailed mechanistic characterization of MGAT5 will aid inhibitor development to correct cancer-associated N-glycosylation

Comprehensive Geriatric Assessment on an Acute Medical Unit: a Qualitative Study of Older People’s and Informal Carer’s Perspectives of the Care and Treatment Received

Objective: This qualitative study was imbedded in a randomized controlled trial evaluating the addition of geriatricians to usual care to enable the comprehensive geriatric assessment process with older patients on acute medical units. The qualitative study explored the perspectives of intervention participants on their care and treatment.Design: A constructivist study incorporating semi-structured interviews that were conducted in patients’ homes within six weeks of discharge from the acute medical unit. These interviews were recorded, transcribed, and analysed using thematic analysis.Setting: An acute medical unit in the United Kingdom.Participants: Older patients (n = 18) and their informal carers (n = 6) discharged directly home from an acute medical unit, who had been in the intervention group of the randomized controlled trial.Results: Three core themes were constructed: (1) perceived lack of treatment on the acute medical unit; (2) nebulous grasp of the role of the geriatrician; and (3) on-going health and activities of daily living needs postdischarge. These needs impacted upon the informal carers, who either took over, or helped the patients to complete their activities of daily living. Despite the help received with activities of daily living, a lot of the patients voiced a desire to complete these activities themselves.Conclusions:The participants perceived they were just monitored and observed on the acute medical unit, rather than receiving active treatment, and spoke of on-going unresolved health and activity of daily living needs following discharge, despite receiving the additional intervention of a geriatrician

Assessment of Exploration Exercise Concepts and Operations Methodology in a BAA Habitat Test Environment

No abstract availabl

Solution structure of the SGTA dimerisation domain and investigation of its interactions with the ubiquitin-like domains of BAG6 and UBL4A

BACKGROUND: The BAG6 complex resides in the cytosol and acts as a sorting point to target diverse hydrophobic protein substrates along their appropriate paths, including proteasomal degradation and ER membrane insertion. Composed of a trimeric complex of BAG6, TRC35 and UBL4A, the BAG6 complex is closely associated with SGTA, a co-chaperone from which it can obtain hydrophobic substrates. METHODOLOGY AND PRINCIPAL FINDINGS: SGTA consists of an N-terminal dimerisation domain (SGTA_NT), a central tetratricopeptide repeat (TPR) domain, and a glutamine rich region towards the C-terminus. Here we solve a solution structure of the SGTA dimerisation domain and use biophysical techniques to investigate its interaction with two different UBL domains from the BAG6 complex. The SGTA_NT structure is a dimer with a tight hydrophobic interface connecting two sets of four alpha helices. Using a combination of NMR chemical shift perturbation, isothermal titration calorimetry (ITC) and microscale thermophoresis (MST) experiments we have biochemically characterised the interactions of SGTA with components of the BAG6 complex, the ubiquitin-like domain (UBL) containing proteins UBL4A and BAG6. We demonstrate that the UBL domains from UBL4A and BAG6 directly compete for binding to SGTA at the same site. Using a combination of structural and interaction data we have implemented the HADDOCK protein-protein interaction docking tool to generate models of the SGTA-UBL complexes. SIGNIFICANCE: This atomic level information contributes to our understanding of the way in which hydrophobic proteins have their fate decided by the collaboration between SGTA and the BAG6 complex

Surfactant Protein-D is essential for immunity to Helminth Infection

Author Summary Infections by parasitic worms are very common, and controlling them is a major medical and veterinary challenge. Very few drugs exist to treat them, and the parasites can develop resistance to these. In order to find new ways to control worm infections, understanding how our immune system responds to them is essential. Many important parasitic worm infections move through the host lung. In this study we show that a major secreted protein in the lung, Surfactant Protein D (SP-D), is essential for immunity to a parasitic worm infection. We found that this protein binds to worm larvae in the lung to help the immune system kill them. Infecting mice that do not express SP-D with worms demonstrates SP-D is important in this immune response. These mice are unable to launch an effective anti-worm immune response and have many more worms in their intestine compared to mice that do express SP-D. We also show that if we increase SP-D levels in the lung the mouse has better immunity to worms. Together this shows for the first time that SP-D is very important for immunity to worm infections

Peptide transport in Bacillus subtilis – structure and specificity in the extracellular solute binding proteins OppA and DppE

Peptide transporters play important nutritional and cell signalling roles in Bacillus subtilis, which are pronounced during stationary phase adaptations and development. Three high-affinity ATP-binding cassette (ABC) family transporters are involved in peptide uptake – the oligopeptide permease (Opp), another peptide permease (App) and a less well-characterized dipeptide permease (Dpp). Here we report crystal structures of the extracellular substrate binding proteins, OppA and DppE, which serve the Opp and Dpp systems, respectively. The structure of OppA was determined in complex with endogenous peptides, mod-elled as Ser-Asn-Ser-Ser, and with the sporulation-promoting peptide Ser-Arg-Asn-Val-Thr, which bind with Kd values of 0.4 and 2 µM, respectively, as measured by isothermal titration calorimetry. Differential scanning fluorescence experiments with a wider panel of ligands showed that OppA has highest affinity for tetra-and penta-peptides. The structure of DppE revealed the unexpected presence of a murein tripeptide (MTP) ligand, l-Ala-d-Glu-meso-DAP, in the peptide binding groove. The mode of MTP binding in DppE is different to that observed in the murein peptide binding protein, MppA, from Escherichia coli, suggesting independent evolution of these proteins from an OppA-like precursor. The presence of MTP in DppE points to a role for Dpp in the uptake and recycling of cell wall peptides, a conclusion that is supported by analysis of the genomic context of dpp, which revealed adjacent genes encoding enzymes involved in muropeptide catabolism in a gene organization that is widely conserved in Firmicutes

Rehabilitation aimed at improving outdoor mobility for people after stroke: a multicentre randomised controlled study (the Getting out of the House Study)

Background: One-third of stroke patients are dependent on others to get outside their homes. This can cause people to become housebound, leading to increased immobility, poor health, isolation and misery. There is some evidence that outdoor mobility rehabilitation can reduce these limitations. Objective: To test the clinical effectiveness and cost-effectiveness of an outdoor mobility rehabilitation intervention for stroke patients. Design: Multicentre, parallel-group randomised controlled trial, with two groups allocated at a 1 : 1 ratio plus qualitative participant interviews. Setting: Fifteen UK NHS stroke services throughout England, Scotland and Wales. Participants: A total of 568 stroke patients who wished to get out of the house more often, mean age of 71 years: 508 reached the 6-month follow-up and 10 were interviewed. Intervention: Control was delivered prior to randomisation to all participants, and consisted of verbal advice and transport and outdoor mobility leaflets. Intervention was a targeted outdoor mobility rehabilitation programme delivered by 29 NHS therapists to 287 randomly chosen participants for up to 12 sessions over 4 months. Main outcome measures: Primary outcome was participant health-related quality of life, measured by the Short Form questionnaire-36 items, version 2 (Social Function domain), 6 months after baseline. Secondary outcomes were functional ability, mobility, number of journeys (from monthly travel diaries), satisfaction with outdoor mobility (SWOM), psychological well-being and resource use [health care and Personal Social Services (PSS)] 6 months after baseline. Carer well-being was recorded. All outcome measures were collected by post and repeated 12 months after baseline. Outcomes for the groups were compared using statistical significance testing and adjusted for multiple membership to account for the effect of multiple therapists at different sites. Interviews were analysed using interpretive phenomenology to explore confidence. Results: A median of seven intervention sessions [interquartile range (IQR) 3–7 sessions], median duration of 369 minutes (IQR 170–691.5 minutes) per participant was delivered. There was no significant difference between the groups on health-related quality of life (social function). There were no significant differences between groups in functional ability, psychological well-being or SWOM at 6- or 12-month follow-ups. There was a significant difference observed for travel journeys with the intervention group being 42% more likely to make a journey compared with the control group [rate ratio 1.42, 95% confidence interval (95% CI) 1.14 to 1.67] at 6 months and 76% more likely (rate ratio 1.76, 95% CI 1.36 to 1.95) at 12 months. The number of journeys was affected by the therapist effect. The mean incremental cost (total NHS and PSS cost) of the intervention was £3413.75 (95% CI –£448.43 to £7121.00), with an incremental quality-adjusted life-year gain of –0.027 (95% CI –0.060 to 0.007) according to the European Quality of Life-5 Dimensions and –0.003 (95% CI –0.016 to 0.006) according to the Short Form questionnaire-6 Dimensions. At baseline, 259 out of 281 (92.2%) participants in the control group were dissatisfied with outdoor mobility but at the 6-month assessment this had reduced to 77.7% (181/233), a 15% reduction. The corresponding reduction in the intervention group was slightly greater (21%) than 268 out of 287 (93.4%) participants dissatisfied with outdoor mobility at baseline to 189 out of 261 (72.4%) at 6 months. Participants described losing confidence after stroke as being detrimental to outdoor mobility. Recruitment and retention rates were high. The intervention was deliverable by the NHS but had a neutral effect in all areas apart from potentially increasing the number of journeys. This was dependent on the therapist effect, meaning that some therapists were more successful than others. The control appeared to affect change. Conclusions: The outdoor mobility intervention provided in this study to these stroke patients was not clinically effective or cost-effective. However, the provision of personalised information and monthly diaries should be considered for all people who wish to get out more

Occurrence of L-iduronic acid and putative D-glucuronyl C5-epimerases in prokaryotes

Glycosaminoglycans (GAGs) are polysaccharides that are typically present in a wide diversity of animal tissue. Most common GAGs are well-characterized and pharmaceutical applications exist for many of these compounds, e.g. heparin and hyaluronan. In addition, also bacterial glycosaminoglycan-like structures exist. Some of these bacterial GAGs have been characterized, but until now no bacterial GAG has been found that possesses the modifications that are characteristic for many of the animal GAGs such as sulfation and C5-epimerization. Nevertheless, the latter conversion may also occur in bacterial and archaeal GAGs, as some prokaryotic polysaccharides have been demonstrated to contain L-iduronic acid. However, experimental evidence for the enzymatic synthesis of L-iduronic acid in prokaryotes is as yet lacking. We therefore performed an in silico screen for D-glucuronyl C5-epimerases in prokaryotes. Multiple candidate C5-epimerases were found, suggesting that many more microorganisms are likely to exist possessing an L-iduronic acid residue as constituent of their cell wall polysaccharides

Generation and transmission of interlineage recombinants in the SARS-CoV-2 pandemic.

We present evidence for multiple independent origins of recombinant SARS-CoV-2 viruses sampled from late 2020 and early 2021 in the United Kingdom. Their genomes carry single-nucleotide polymorphisms and deletions that are characteristic of the B.1.1.7 variant of concern but lack the full complement of lineage-defining mutations. Instead, the remainder of their genomes share contiguous genetic variation with non-B.1.1.7 viruses circulating in the same geographic area at the same time as the recombinants. In four instances, there was evidence for onward transmission of a recombinant-origin virus, including one transmission cluster of 45 sequenced cases over the course of 2 months. The inferred genomic locations of recombination breakpoints suggest that every community-transmitted recombinant virus inherited its spike region from a B.1.1.7 parental virus, consistent with a transmission advantage for B.1.1.7's set of mutations.The COG-UK Consortium is supported by funding from the Medical Research Council (MRC) part of UK Research & Innovation (UKRI), the National Institute of Health Research (NIHR) (MC_PC_19027), and Genome Research Limited, operating as the Wellcome Sanger Institute. O.G.P. was supported by the Oxford Martin School. J.T.M., R.M.C., N.J.L., and A.R. acknowledge the support of the Wellcome Trust (Collaborators Award 206298/Z/17/Z – ARTIC network). D.L.R. acknowledges the support of the MRC (MC_UU_12014/12) and the Wellcome Trust (220977/Z/20/Z). E.S. and A.R. are supported by the European Research Council (grant agreement no. 725422 – ReservoirDOCS). T.R.C. and N.J.L. acknowledge the support of the MRC, which provided the funding for the MRC CLIMB infrastructure used to analyze, store, and share the UK sequencing dataset (MR/L015080/1 and MR/T030062/1). The samples sequenced in Wales were sequenced partly using funding provided by the Welsh Government

Evaluation of the guide to action care home fall prevention programme in care homes for older people: A multi-centre, single blinded, cluster randomised controlled trial (FINCH)

BackgroundFalls in care home residents are common, unpleasant, costly and difficult to prevent. Trial DesignThe objective was to evaluate the clinical and cost effectiveness of the Guide to Action for Falls Prevention Care Homes, GtACH) in which care home staff were trained and supported in the systematic use of a multi-domain decision support tool and identify issues affecting subsequent implementation. A two-arm parallel design, multi-centre, cluster randomised controlled trial of the GtACH programme and usual falls prevention in older care home residents was conducted with embedded process evaluation and economic evaluation. MethodThe study was conducted in care homes from ten UK sites. The primary trial outcome was the rate of falls per resident participant occurring during the 90-day period between 91 days and 180 days post-randomisation. The primary outcome for the cost effectiveness analysis was the cost per fall averted and for the cost utility analysis was the incremental cost per QALY. Secondary outcomes included the rate of falls over days 0-90 and 181-360 post randomisation, activity levels, dependency, and fractures. Care homes were randomised on a 1:1 basis to the GtACH programme or usual care, via a secure web-based randomisation service. Research assistants (RAs), resident participants and staff informants were blind to allocation at recruitment. RAs were blind to allocation at follow up. Data from NHS Digital were extracted blindly. The number of falls per resident was compared between groups using a negative binomial regression model (GEE).Results84 care homes were randomised, 39 to GtACH and 45 to usual care. 1657 residents consented and provided baseline measures, mean age 85 years, 32% men. GtACH training was delivered to 1051 staff (71% of eligible staff) over 146 group sessions. Primary RCT outcome data were available for 630 of the GtACH participants and 712 usual care participants. The primary RCT outcome result showed an unadjusted Incidence Rate Ratio (IRR) of 0.57 (95% CI 0.45-0.71, p<0.01) in favour of the GtACH programme. Fall rates were also lower in the GtACH group in the period 0-90 days, but there were no other differences between groups in the secondary outcomes. Care home staff valued the training, the systematic strategies and the specialist peer support, but there was limited incorporation of the GtACH documentation into routine care home practice. No adverse events were recorded. The incremental cost per DEMQoL-based QALY was £20,889.42 and £4,543.69 per EQ-5D based QALY. Mean falls were 1.889 (sd 3.662) in the GtACH arm and 2.747 (sd 7.414) in the usual care arm. Therefore, 0.858 falls were averted. The base case incremental cost per fall averted was £190.62ConclusionThe GtACH programme significantly reduced the rate of falls in the study care homes, without restricting residents’ activity levels or increasing their dependency and was cost effective at current thresholds in the UK NHS. Widespread implementation of the programme is justified. Trial registrationTrial registration number: ISRCTN34353836. Protocol V6 14 November 2017Funding DetailsThe National Institute for Health Research (NIHR) HTA programm