Binding pocket stabilization by high-throughput screening of yeast display libraries

Protein dynamics have a great influence on the binding pockets of some therapeutic targets. Flexible protein binding sites can result in transient binding pocket formation which might have a negative impact on drug screening efforts. Here, we describe a protein engineering strategy with FK506-binding protein 51 (FKBP51) as a model protein, which is a promising target for stress-related disorders. High-throughput screening of yeast display libraries of FKBP51 resulted in the identification of variants exhibiting higher affinity binding of conformation-specific FKBP51 selective inhibitors. The gene libraries of a random mutagenesis and site saturation mutagenesis of the FK1 domain of FKBP51 encoding sequence were used to create a yeast surface display library. Fluorescence-activated cell sorting for FKBP51 variants that bind conformation-specific fluorescently labeled ligands with high affinity allowed for the identification of 15 different protein variants with improved binding to either, or both FKBP51-specific ligands used in the screening, with improved affinities up to 34-fold compared to the wild type. These variants will pave the way to a better understanding of the conformational flexibility of the FKBP51 binding pocket and may enable the isolation of new selective ligands that preferably and selectively bind the active site of the protein in its open conformation state

Nurses' perceptions of aids and obstacles to the provision of optimal end of life care in ICU

Contains fulltext : 172380.pdf (publisher's version ) (Open Access

Mortality from gastrointestinal congenital anomalies at 264 hospitals in 74 low-income, middle-income, and high-income countries: a multicentre, international, prospective cohort study

Summary Background Congenital anomalies are the fifth leading cause of mortality in children younger than 5 years globally. Many gastrointestinal congenital anomalies are fatal without timely access to neonatal surgical care, but few studies have been done on these conditions in low-income and middle-income countries (LMICs). We compared outcomes of the seven most common gastrointestinal congenital anomalies in low-income, middle-income, and high-income countries globally, and identified factors associated with mortality. Methods We did a multicentre, international prospective cohort study of patients younger than 16 years, presenting to hospital for the first time with oesophageal atresia, congenital diaphragmatic hernia, intestinal atresia, gastroschisis, exomphalos, anorectal malformation, and Hirschsprung’s disease. Recruitment was of consecutive patients for a minimum of 1 month between October, 2018, and April, 2019. We collected data on patient demographics, clinical status, interventions, and outcomes using the REDCap platform. Patients were followed up for 30 days after primary intervention, or 30 days after admission if they did not receive an intervention. The primary outcome was all-cause, in-hospital mortality for all conditions combined and each condition individually, stratified by country income status. We did a complete case analysis. Findings We included 3849 patients with 3975 study conditions (560 with oesophageal atresia, 448 with congenital diaphragmatic hernia, 681 with intestinal atresia, 453 with gastroschisis, 325 with exomphalos, 991 with anorectal malformation, and 517 with Hirschsprung’s disease) from 264 hospitals (89 in high-income countries, 166 in middleincome countries, and nine in low-income countries) in 74 countries. Of the 3849 patients, 2231 (58·0%) were male. Median gestational age at birth was 38 weeks (IQR 36–39) and median bodyweight at presentation was 2·8 kg (2·3–3·3). Mortality among all patients was 37 (39·8%) of 93 in low-income countries, 583 (20·4%) of 2860 in middle-income countries, and 50 (5·6%) of 896 in high-income countries (p<0·0001 between all country income groups). Gastroschisis had the greatest difference in mortality between country income strata (nine [90·0%] of ten in lowincome countries, 97 [31·9%] of 304 in middle-income countries, and two [1·4%] of 139 in high-income countries; p≤0·0001 between all country income groups). Factors significantly associated with higher mortality for all patients combined included country income status (low-income vs high-income countries, risk ratio 2·78 [95% CI 1·88–4·11], p<0·0001; middle-income vs high-income countries, 2·11 [1·59–2·79], p<0·0001), sepsis at presentation (1·20 [1·04–1·40], p=0·016), higher American Society of Anesthesiologists (ASA) score at primary intervention (ASA 4–5 vs ASA 1–2, 1·82 [1·40–2·35], p<0·0001; ASA 3 vs ASA 1–2, 1·58, [1·30–1·92], p<0·0001]), surgical safety checklist not used (1·39 [1·02–1·90], p=0·035), and ventilation or parenteral nutrition unavailable when needed (ventilation 1·96, [1·41–2·71], p=0·0001; parenteral nutrition 1·35, [1·05–1·74], p=0·018). Administration of parenteral nutrition (0·61, [0·47–0·79], p=0·0002) and use of a peripherally inserted central catheter (0·65 [0·50–0·86], p=0·0024) or percutaneous central line (0·69 [0·48–1·00], p=0·049) were associated with lower mortality. Interpretation Unacceptable differences in mortality exist for gastrointestinal congenital anomalies between lowincome, middle-income, and high-income countries. Improving access to quality neonatal surgical care in LMICs will be vital to achieve Sustainable Development Goal 3.2 of ending preventable deaths in neonates and children younger than 5 years by 2030

Table1_Binding pocket stabilization by high-throughput screening of yeast display libraries.DOCX

Protein dynamics have a great influence on the binding pockets of some therapeutic targets. Flexible protein binding sites can result in transient binding pocket formation which might have a negative impact on drug screening efforts. Here, we describe a protein engineering strategy with FK506-binding protein 51 (FKBP51) as a model protein, which is a promising target for stress-related disorders. High-throughput screening of yeast display libraries of FKBP51 resulted in the identification of variants exhibiting higher affinity binding of conformation-specific FKBP51 selective inhibitors. The gene libraries of a random mutagenesis and site saturation mutagenesis of the FK1 domain of FKBP51 encoding sequence were used to create a yeast surface display library. Fluorescence-activated cell sorting for FKBP51 variants that bind conformation-specific fluorescently labeled ligands with high affinity allowed for the identification of 15 different protein variants with improved binding to either, or both FKBP51-specific ligands used in the screening, with improved affinities up to 34-fold compared to the wild type. These variants will pave the way to a better understanding of the conformational flexibility of the FKBP51 binding pocket and may enable the isolation of new selective ligands that preferably and selectively bind the active site of the protein in its open conformation state.</p

Prediction of Patient Satisfaction after Treatment of Chronic Neck Pain with Mulligan’s Mobilization

Chronic neck pain is among the most common types of musculoskeletal pain. Manual therapy has been shown to have positive effects on this type of pain, but there are not yet many predictive models for determining how best to apply manual therapy to the different subtypes of neck pain. The aim of this study is to develop a predictive learning approach to determine which basal outcome could give a prognostic value (Global Rating of Change, GRoC scale) for Mulligan’s mobilization technique and to identify the most important predictive factors for recovery in chronic neck pain subjects in four key areas: the number of treatments, time of treatment, reduction of pain, and range of motion (ROM) increase. A prospective cohort dataset of 80 participants with chronic neck pain diagnosed by their family doctor was analyzed. Logistic regression and machine learning modeling techniques (Generalized Boosted Models, Support Vector Machine, Kernel, Classsification and Decision Trees, Random Forest and Neural Networks) were each used to form a prognostic model for each of the nine outcomes obtained before and after intervention: disability—neck disability index (NDI), patient satisfaction (GRoC), quality of life (12-Item Short Form Survey, SF-12), State-Trait Anxiety Inventory (STAI), Beck Depression Inventory (BDI II), pain catastrophizing scale (ECD), kinesiophobia-Tampa scale of kinesiophobia (TSK-11), Pain Intensity Visual Analogue Scale (VAS), and cervical ROM. Pain descriptions from the subjects and pain body diagrams guided the physical examination. The most important predictive factors for recovery in chronic neck pain patients indicated that the more anxiety and the lower the ROM of lateroflexion, the higher the probability of success with the Mulligan concept treatment

Antioxidants in frying: Analysis and evaluation of efficacy

The action of antioxidants is well known under room temperature conditions during storage or at the moderate temperatures of the accelerated tests used to measure the oil stability. However, efficacy of antioxidants under frying conditions is far more difficult to be evaluated and defined because availability of air is lower and variable, and both oxidation and thermal reactions are simultaneously involved. This review is focused on the analysis and evaluation of efficacy of antioxidants in frying. Specific aspects of the action of natural and synthetic antioxidants at high temperature are discussed, and the most important methods used for the analysis of antioxidants and their efficacy are described.Peer Reviewe

2do Workshop Internacional de Investigación formativa en ciencias para afrontar tiempos de cambio

La presente propuesta de investigación formativa en el contexto de la didáctica de las ciencias naturales, matemáticas y tecnología, tiene el objetivo de convertirse en una estrategia didáctica que desarrollará las capacidades científicas y entrará en tensión con las competencias exigidas por el Ministerio de Educación Nacional (MEN) para la presentación de la prueba saber 11, esta misma profundizará en la búsqueda de respuestas frente al desinterés evidente por el estudio de las ciencias en estudiantes cursando ciclo V de instituciones educativas distritales. Para ello realizaremos una prueba piloto con estudiantes del ciclo V del colegio Japón IED, más específicamente con la participación de aquellos alumnos que cuenten con capacidades y aptitudes científicas para conformar y ser parte del semillero “EduQVersa Junior” del mismo colegio. Lo anterior con el fin de obtener resultados cualitativos y cuantitativos para posteriormente organizar una feria científica, ésta misma pretenderá potencializar el interés y asombro en el estudio de las ciencias por medio de proyectos basados con la metodología STEAM, para ulteriormente ser expuesto el día del campo científico, el Workshop, entre otros eventos de difusión de índole local, nacional e internacional. Con los anteriores objetivos se espera fomentar procesos investigativos iniciales en el campo de las ciencias y buscarán incentivar en general en los estudiantes de ciclo V un mayor interés y continuidad en la formación de su educación superior en el estudio de esta área, siendo una apuesta de focalización que se entrecruza con el componente académico y la consolidación del proyecto de

ESICM LIVES 2016: part two : Milan, Italy. 1-5 October 2016.

Meeting abstrac