Activation in the family of Candida rugosa isolipases by polyethylene glycol

Disponible en: http://www.xtal.iqfr.csic.es/publications/jmolcat2005.pdfWe have investigated activation of two isoenzymes (lip1 and lip3) from Candida rugosa in polyethylene glycol (PEG) media. Aqueous solutions of PEG 8000 and 20,000 activate lip3 but not lip1 from C. rugosa. Maximum activation (260%) of lip3 requires 6 h of pre-incubation with PEG 8000 (4%, w/v). PEG seems to shift the equilibrium between the open and the closed forms of lip3 towards the active conformation. Inhibition experiments demonstrate that ligands have easier access to the lip3 active site than to the lip1 active site, both in the presence and the absence of PEG. The presence of PEG in the crystallization medium is responsible for reported differences in the crystal structures of lip1 and lip3. A comparative analysis of crystallographic models of lip1 and lip3 suggests a role for PEG in activation of lip3 and further stabilization of the activated/open form via dimerization in aqueous media.This work has been financed by the Spanish CICYT (PB92/0495 and BIO 96/0837). A predoctoral fellowship for M.F.P. was provided by the Autonomous Community of Madrid (Spain).Peer reviewe

The structure of the C-terminal domain of the protein kinase AtSOS2 bound to the calcium sensor AtSOS3

Disponible en: http://www.xtal.iqfr.csic.es/publications/molcell2007.pdfThe plant SOS2 family of protein kinases and their interacting activators, the SOS3 family of calcium-binding proteins, function together in decoding calcium signals elicited by different environmental stimuli. SOS2 is activated by Ca-SOS3 and subsequently phosphorylates the ion transporter SOS1 to bring about cellular ion homeostasis under salt stress. In addition to possessing the kinase activity, members of the SOS2 family of protein kinases can bind to protein phosphatase 2Cs. The crystal structure of the binary complex of Ca-SOS3 with the C-terminal regulatory moiety of SOS2 resolves central questions regarding the dual function of SOS2 as a kinase and a phosphatase-binding protein. A comparison with the structure of unbound SOS3 reveals the basis of the molecular function of this family of kinases and their interacting calcium sensors. Furthermore, our study suggests that the structure of the phosphataseinteraction domain of SOS2 defines a scaffold module conserved from yeast to human.Was supported by a Formacion de Personal Investigador studentship from Ministerio de Educación, Cultura y Deporte, and a Postgraduate I3P Fellowship from the Consejo Superior de Investigaciones Cientificas. A.A. thanks the European Synchrotron Radiation Facility (ESRF) for access to the synchrotron radiation source. This work was funded by the BFU2005-06388-C04-02/BMC grant of the Spanish ‘‘Plan Nacional’’ (MEC) and ‘‘Factoría de Cristalización’’ Consolider Ingenio 2010 to A.A. and by U.S. National Institutes of Health grant R01GM59138 to J.-K.Z.Peer reviewe

Crystal structures of bacterial peptidoglycan amidase AmpD and an unprecedented activation mechanism

9 pags, 5 figs, 2 tabsAmpD is a cytoplasmic peptidoglycan (PG) amidase involved in bacterial cell-wall recycling and in induction of β-lactamase, a key enzyme of β-lactam antibiotic resistance. AmpD belongs to the amidase-2 family that includes zinc-dependent amidases and the peptidoglycan-recognition proteins (PGRPs), highly conserved pattern-recognition molecules of the immune system. Crystal structures of Citrobacter freundii AmpD were solved in this study for the apoenzyme, for the holoenzyme at two different pH values, and for the complex with the reaction products, providing insights into the PG recognition and the catalytic process. These structures are significantly different compared with the previously reported NMR structure for the same protein. TheNMRstructure does not possess an accessible active site and shows the protein in what is proposed herein as an inactive "closed" conformation. The transition of the protein from this inactive conformation to the active "open" conformation, as seen in the x-ray structures, was studied by targeted molecular dynamics simulations, which revealed large conformational rearrangements (as much as 17 Å ) in four specific regions representing one-third of the entire protein. It is proposed that the large conformational change that would take the inactive NMR structure to the active x-ray structure represents an unprecedented mechanism for activation of AmpD. Analysis is presented to argue that this activation mechanism might be representative of a regulatory process for other intracellular members of the bacterial amidase-2 family of enzymes. © 2011 by The American Society for Biochemistry and Molecular Biology, Inc.This work was supported, in whole or in part, by the National Institutes of Health. This work was also supported by grants from the Spanish Ministry of Science and Technology (BFU2008-01711), EU-CP223111 (CARE-PNEUMO, European Union), and the COMBACT program (S-BIO-0260/2006). We acknowledge the Spanish Ministerio de Ciencia e Innovación (PI201060E013) and Consejo Superior de Investigaciones Científicas for financial support and for provision of synchrotron radiation facilitie

Insights into pneumococcal fratricide from the crystal structures of the modular killing factor LytC

7 pags, 3 figs, 1 tabThe first structure of a pneumococcal autolysin, that of the LytC lysozyme, has been solved in ternary complex with choline and a pneumococcal peptidoglycan (PG) fragment. The active site of the hydrolase module is not fully exposed but is oriented toward the choline-binding module, which accounts for its unique in vivo features in PG hydrolysis, its activation and its regulatory mechanisms. Because of the unusual hook-shaped conformation of the multimodular protein, it is only able to hydrolyze non-cross-linked PG chains, an assertion validated by additional experiments. These results explain the activation of LytC by choline-binding protein D (CbpD) in fratricide, a competence-programmed mechanism of predation of noncompetent sister cells. The results provide the first structural insights to our knowledge into the critical and central function that LytC plays in pneumococcal virulence and explain a long-standing puzzle of how murein hydrolases can be controlled to avoid self-lysis during bacterial growth and division. © 2010 Nature America, Inc. All rights reserved.This work was supported by grants from the Spanish Ministry of Science and Technology (BFU2008-01711, SAF2006-00390), EU-CP223111 (CAREPNEUMO, European Union), the COMBACT program (S-BIO-0260/2006) and CIBER de Enfermedades Respiratorias (CIBERES). CIBERES is an initiative of Instituto de Salud Carlos III. The work in the US was supported by the US National Institutes of Health. W.V. was supported by the European Commission (EUR-INTAFAR project). I.P.-D. was a fellow of the Consejo Superior de Investigaciones Científicas

Augmentation of EMDR with multifocal transcranial current stimulation (MtCS) in the treatment of fibromyalgia : study protocol of a double-blind randomized controlled exploratory and pragmatic trial

Fibromyalgia (FM) is a generalized, widespread chronic pain disorder affecting 2.7% of the general population. In recent years, different studies have observed a strong association between FM and psychological trauma. Therefore, a trauma-focused psychotherapy, such as eye movement desensitization and reprocessing (EMDR), combined with a non-invasive brain stimulation technique, such as multifocal transcranial current stimulation (MtCS), could be an innovative adjunctive treatment option. This double-blind randomized controlled trial (RCT) analyzes if EMDR therapy is effective in the reduction of pain symptoms in FM patients and if its potential is boosted with the addition of MtCS. Forty-five patients with FM and a history of traumatic events will be randomly allocated to Waiting List, EMDR + active-MtCS, or EMDR + sham-MtCS. Therapists and patients will be kept blind to MtCS conditions, and raters will be kept blind to both EMDR and MtCS. All patients will be evaluated at baseline, post-treatment, and follow-up at 6 months after post-treatment. Evaluations will assess the following variables: sociodemographic data, pain, psychological trauma, sleep disturbance, anxiety and affective symptoms, and wellbeing. This study will provide evidence of whether EMDR therapy is effective in reducing pain symptoms in FM patients, and whether the effect of EMDR can be enhanced by MtCS. ClinicalTrials.gov . Registered on 2 August 2019. The online version contains supplementary material available at 10.1186/s13063-021-05042-w

Strain Effects in Protonated Carbonyl Compounds. An Experimental and ab Initio Treatment of Acyclic Carboxamides and Ketones

Strain effects have been quantitatively evaluated for a set of 22 compounds including ketones (R2CO), carboxamides (RCONH2), and N,N-dimethylcarboxamides (RCONMe2), where R = Me, Et, i-Pr, t-Bu, 1-adamantyl (1-Ad), in their neutral and protonated forms. To this end, use was made of the gas-phase proton affinities and standard enthalpies of formation of these compounds in the gas phase, as determined by Fourier transform ion cyclotron resonance mass spectrometry (FT ICR) and thermochemical techniques, respectively. The structures of 1-AdCOMe and (1-Ad)2CO were determined by X-ray crystallography. Quantum-mechanical calculations, at levels ranging from AM1 to MP2/6-311+G(d,p)//6-31G(d), were performed on the various neutral and protonated species. Constrained space orbital variation (CSOV) calculations were carried out on selected protonated species to further assess the contributions of the various stabilizing factors. Taking neutral and protonated methyl ketones as references, we constructed isodesmic reactions that provided, seemingly for the first time, quantitative measures of strain in the protonated species. A combination of these data with the results of theoretical calculations (which also included several “computational experiments”) lead to a unified, conceptually satisfactory, quantitative description of these effects and their physical link to structural properties of the neutral and protonated species.This work was supported by grants PB 93-0289-C02 and PB-93-0142-C03-01 from the Spanish D.G.I.C.Y.T. Work by H.H. was supported by the Moroccan Ministry of Education and C.S.I.C

Improving the users' balance with video-games

[EN] In recent years, the computer and video-games industry has created new home-based, low-cost technologies for capturing human movements, which make the combination of video-games and biomechanical analysis a feasible reality. The iStoppFalls project takes advantage of this opportunity for assessing the risk of falling and improve the users¿ balance during their interaction with exercise games. One of the challenges is achieving measures of enough quality, from a system that is originally designed for capturing gross movements. This problem is solved by a redefinition of the measurement protocols, plus advanced analysis of the variables captured by the videogame sensors, in order to reduce the distance between required and achieved precision. Validated fall risk assessment criteria have been adapted to exercises that can be made at home, without supervision or additional instrumentation. A skeleton model with 22 degrees of freedom has been tracked by Kinect, with joint angles corrected by a particle filter.[ES] En los últimos años, la industria Mejorar el equilibrio con videojuegos informática y de videojuegos ha creado nuevas tecnologías domésticas y de bajo coste para la captura de movimientos humanos, lo cual abre nuevas posibilidades de combinar videojuegos y análisis biomecánicos. El proyecto iStoppFalls, financiado por el 7º Programa Marco de la Unión Europea, aprovecha esta oportunidad para evaluar el riesgo de caídas durante la interacción de los usuarios al realizar ejercicios en videojuegos y llevar a cabo programas de entrenamiento para mejorar el equilibrio. Uno de los retos consiste en conseguir medidas de suficiente calidad, a partir de un sistema originalmente diseñado para capturar movimientos poco precisos. Este problema se resuelve redefiniendo los protocolos de medida y realizando un análisis avanzado de las variables capturadas por los sensores de los videojuegos. Para ello se han adaptado criterios validados de evaluación del riesgo de caída, convirtiéndolos en ejercicios y juegos que pueden realizarse en el hogar, sin supervisión ni instrumentación. El movimiento realizado durante estos ejercicios se monitoriza a través del sensor Kinect de Microsoft, utilizando un modelo esquelético de 22 grados de libertad, con ángulos articulares corregidos mediante un filtro de partículas.iStoppFalls (FP7-ICT-2011-7- 287361) es un proyecto financiado por el 7º Programa Marco de la Unión Europea.De Rosario Martínez, H.; Avila Carrasco, C.; Fos Ros, F.; Medina Ripoll, E.; Castellanos Hernández, WE.; Bollain Pastor, C.; Poveda Puente, R.... (2013). Mejorar el equilibrio con videojuegos. Revista de biomecánica. 59:31-34. http://hdl.handle.net/10251/38684S31345

Evaluation of turbulent dissipation rate retrievals from Doppler Cloud Radar

Turbulent dissipation rate retrievals from cloud radar Doppler velocity measurements are evaluated using independent, in situ observations in Arctic stratocumulus clouds. In situ validation data sets of dissipation rate are derived using sonic anemometer measurements from a tethered balloon and high frequency pressure variation observations from a research aircraft, both flown in proximity to stationary, ground-based radars. Modest biases are found among the data sets in particularly low- or high-turbulence regimes, but in general the radar-retrieved values correspond well with the in situ measurements. Root mean square differences are typically a factor of 4-6 relative to any given magnitude of dissipation rate. These differences are no larger than those found when comparing dissipation rates computed from tetheredballoon and meteorological tower-mounted sonic anemometer measurements made at spatial distances of a few hundred meters. Temporal lag analyses suggest that approximately half of the observed differences are due to spatial sampling considerations, such that the anticipated radar-based retrieval uncertainty is on the order of a factor of 2-3. Moreover, radar retrievals are clearly able to capture the vertical dissipation rate structure observed by the in situ sensors, while offering substantially more information on the time variability of turbulence profiles. Together these evaluations indicate that radar-based retrievals can, at a minimum, be used to determine the vertical structure of turbulence in Arctic stratocumulus clouds