Thermophile-specific proteins: the gene product of aq_1292 from Aquifex aeolicus is an NTPase

BACKGROUND: To identify thermophile-specific proteins, we performed phylogenetic patterns searches of 66 completely sequenced microbial genomes. This analysis revealed a cluster of orthologous groups (COG1618) which contains a protein from every thermophile and no sequence from 52 out of 53 mesophilic genomes. Thus, COG1618 proteins belong to the group of thermophile-specific proteins (THEPs) and therefore we here designate COG1618 proteins as THEP1s. Since no THEP1 had been analyzed biochemically thus far, we characterized the gene product of aq_1292 which is THEP1 from the hyperthermophilic bacterium Aquifex aeolicus (aaTHEP1). RESULTS: aaTHEP1 was cloned in E. coli, expressed and purified to homogeneity. At a temperature optimum between 70 and 80°C, aaTHEP1 shows enzymatic activity in hydrolyzing ATP to ADP + P(i )with k(cat )= 5 × 10(-3 )s(-1 )and K(m )= 5.5 × 10(-6 )M. In addition, the enzyme exhibits GTPase activity (k(cat )= 9 × 10(-3 )s(-1 )and K(m)= 45 × 10(-6 )M). aaTHEP1 is inhibited competitively by CTP, UTP, dATP, dGTP, dCTP, and dTTP. As shown by gel filtration, aaTHEP1 in its purified state appears as a monomer. The enzyme is resistant to limited proteolysis suggesting that it consists of a single domain. Although THEP1s are annotated as "predicted nucleotide kinases" we could not confirm such an activity experimentally. CONCLUSION: Since aaTHEP1 is the first member of COG1618 that is characterized biochemically and functional information about one member of a COG may be transferred to the entire COG, we conclude that COG1618 proteins are a family of thermophilic NTPases

Crystal structure of THEP1 from the hyperthermophile Aquifex aeolicus: a variation of the RecA fold

RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are.Abstract Background aaTHEP1, the gene product of aq_1292 from Aquifex aeolicus, shows sequence homology to proteins from most thermophiles, hyperthermophiles, and higher organisms such as man, mouse, and fly. In contrast, there are almost no homologous proteins in mesophilic unicellular microorganisms. aaTHEP1 is a thermophilic enzyme exhibiting both ATPase and GTPase activity in vitro. Although annotated as a nucleotide kinase, such an activity could not be confirmed for aaTHEP1 experimentally and the in vivo function of aaTHEP1 is still unknown. Results Here we report the crystal structure of selenomethionine substituted nucleotide-free aaTHEP1 at 1.4 Å resolution using a multiple anomalous dispersion phasing protocol. The protein is composed of a single domain that belongs to the family of 3-layer (α/β/α)-structures consisting of nine central strands flanked by six helices. The closest structural homologue as determined by DALI is the RecA family. In contrast to the latter proteins, aaTHEP1 possesses an extension of the β-sheet consisting of four additional β-strands. Conclusion We conclude that the structure of aaTHEP1 represents a variation of the RecA fold. Although the catalytic function of aaTHEP1 remains unclear, structural details indicate that it does not belong to the group of GTPases, kinases or adenosyltransferases. A mainly positive electrostatic surface indicates that aaTHEP1 might be a DNA/RNA modifying enzyme. The resolved structure of aaTHEP1 can serve as paradigm for the complete THEP1 family.Published versio

On the cytotoxicity of HCR-NTPase in the neuroblastoma cell line SH-SY5Y

Pasdziernik M, Kaltschmidt B, Kaltschmidt C, Klinger C, Kaufmann M. On the cytotoxicity of HCR-NTPase in the neuroblastoma cell line SH-SY5Y. BMC Research Notes. 2009;2(1):102.Background: The human cancer-related nucleoside triphosphatase (HCR-NTPase) is overexpressed in several tumour tissues including neuroblastoma. HCR-NTPase is an enzyme exhibiting a slow in vitro activity in hydrolysing nucleosidetriphosphates. However, its in vivo function is still unknown. To learn more about the physiological role of HCR-NTPase, we both overexpressed and silenced it in the neuroblastoma cell line SH-SY5Y. Findings: No effect was observed when the expression of endogenously expressed HCR-NTPase in the cells was silenced by RNA interference. On the other hand, overexpression of HCR-NTPase led to cytotoxicity of the protein in SH-SY5Y cells. Even if the catalytic essential amino acid glutamate 114 was replaced by alanine (E114A-HCR-NTPase), the protein remained cytotoxic. The results could be confirmed by successfully rescuing the cells via RNA interference. Conclusion: Although expressed in several tumours, at least in SH-SY5Y, HCR-NTPase is not essential for the cells to survive. Increased levels of the protein lead to cytotoxicity due to physical intracellular interactions rather than hydrolysis of nucleosidetriphosphates by its intrinsic residual enzymatic activity

Web services enabled architecture coupling data and functional resources

Web services are the backbone of WISDOM system, an information and visualisation system supporting decision makers in the fields of water related management processes based on open source technologies. They enable the distributed and loosely coupled, component based architecture of the system. In cooperating OGC compliant web services for data access, visualisation and data processing the system is extendible to external data resources and other proprietary software solutions. The base idea behind the designed and prototypically implemented WISDOM techniques is the orchestration of decoupled web resources representing data sets and functionality to model more complex business processes quite easily. The system covers most aspects of administrative business processes including spatial and non-spatial data ingestion and dissemination, necessary data processing and visualisation techniques. In combination with a semantics enabled data management WISDOM system is capable to produce value added information products to water management related tasks autonomously. These compound data and processing resource chains are implemented to facilitate certain identified business processes in regional administration. Clients like data and information explorers supporting manual interaction as human machine interfaces or automated data access of value adding operations accomplish, respectively trigger these integrative chains. As an example the same data and processing infrastructure is used to visualise data in map clients as WMS or access data as WCS, resp. WFS for further processing which can furthermore trigger additional actions like feeding reports or requesting auxiliary data

Laboratory experiments on the interaction of a buoyant coastal current with a canyon : application to the East Greenland Current

Author Posting. © American Meteorological Society, 2009. This article is posted here by permission of American Meteorological Society for personal use, not for redistribution. The definitive version was published in Journal of Physical Oceanography 39 (2009): 1258-1271, doi:10.1175/2008JPO4028.1.This paper presents a set of laboratory experiments focused on how a buoyant coastal current flowing over a sloping bottom interacts with a canyon and what controls the separation, if any, of the current from the upstream canyon bend. The results show that the separation of a buoyant coastal current depends on the current width W relative to the radius of curvature of the bathymetry ρc. The flow moved across the mouth of the canyon (i.e., separated) for W/ρc > 1, in agreement with previous results. The present study extends previous work by examining both slope-controlled and surface-trapped currents, and using a geometry specific to investigating buoyant current–canyon interaction. The authors find that, although bottom friction is important in setting the position of the buoyant front, the separation process driven by the inertia of the flow could overcome even the strongest bathymetric influence. Application of the laboratory results to the East Greenland Current (EGC), an Arctic-origin buoyant current that is observed to flow in two branches south of Denmark Strait, suggests that the path of the EGC is influenced by the large canyons cutting across the shelf, as the range of W/ρc in the ocean spans those observed in the laboratory. What causes the formation of a two-branched EGC structure downstream of the Kangerdlugssuaq Canyon (68°N, 32°W) is still unclear, but potential mechanisms are discussed.This work was partially funded by NSF Grant OCE-0450658. DS also received support from the Academic Programs Office of the Woods Hole Oceanographic Institution, while CC had partial support from NSF OCE-0350891

Hearing Loss at High Frequencies and Oxidative Stress: A New Paradigm for Different Etiologies

The clinical assessment of hearing loss has been transformed and revised in terms of interpreting the characteristics of patterns found in relation to the relative frequency of certain diseases. However, increasing the threshold to 4 kHz as a starting point for hearing loss has shown to be common to different diseases such as noise-induced hearing loss. In noise-induced hearing loss, for example, six mechanisms can be considered: conversion of sound pressure level into hearing level, vascular failure in the cochlear region responsible for hearing at 4 kHz, sound wave propagation velocity is very high and causes the displacement amplitude in the cochlear duct, the structure anatomy of the cochlea causes a collision of fluids in the first curve of the cochlea, characteristics of auricular pavilion resonance and external auditory canal, and sound attenuation of the acoustic reflex. It is hoped that this new paradigm for the different hearing losses will result in a different approach to the physiological changes that affect the auditory system in the form of high-frequency hearing loss. As such, preventing, treating, and avoiding exacerbations are possibilities to be investigated in order to guarantee efficient communication and quality of life for individuals

Correlation between cholangiopancreatography by magnetic resonance and the endoscopic retrograde colangio pancreatography in hospitalized by biliary lithiasis in the University Hospital San Ignacio (Bogota-Colombia) between 2005 to 2011

Objetivo: Determinar el grado de acuerdo entre la colangiopancreatografía por resonancia magnética (CPRM) y los hallazgos por colangiopancreatografía endoscópica retrograda (CPRE) en pacientes hospitalizados por enfermedad biliar litiásica en un periodo de tiempo en un hospital de referencia. Material y métodos: Tipo de estudio evaluación de prueba diagnóstica retrospectivo, tipo de muestra no probabilístico por conveniencia, el cálculo de tamaño de muestra fue calculado a partir de los datos obtenidos de la prueba piloto y fue de 320 pacientes. Se recolectaron 354 pacientes los cuales se sometieron a ambos estudios (CPRM y CPRE), para evaluar la correlación entre estas 2 pruebas se realizó mediante la variante nominal Kappa. Resultados: cumplieron con criterios de inclusión 354 pacientes, 226 mujeres (63,8%), hombres 128 (36,2%), edad mediana 48 años. Se observó dilatación del colédoco por CRMN en el 60%, sensibilidad del 96,7%, especificidad del 40%, índice de Kappa 0,406 (IC 95%: 0,32-0,50) que indica una concordancia moderada. Para la presencia de cálculo coledociano en CPRM fue 54%, con Sensibilidad del 72,6%, especificidad del 68,3%, índice de Kappa 0,409 (IC 95%: 0,31-0,51) concordancia moderada. Para cálculos en vesícula índice Kappa 0,246 débil. Se identificó 27% de falsos negativos de CRMN para detección de cálculo coledociano. Conclusiones: La CPRM y la CPRE tuvieron una correlación moderada según índice Kappa en la detección de dilatación y cálculos coledocianos en nuestra cohorte de pacientes con probabilidad intermedia. El número de pacientes falsos negativos para coledolitiasis mediante CPRM, plantea la necesidad de búsqueda de otros estudios prospectivos aleatorizados como la ultrasonografía endoscópica biliopancreática que comparen características operativas con CPRM en pacientes de probabilidad intermedia para coledocolitiasis.Q4Artículo original226-230Objetive: Determine the grade of agreement between the colangiopancreatography by magnetic resonance (CPRM) and the endoscopic retrograde colangio pancreatography (ERCP) in hospitalized patients by biliary lithiasis in a period of time in a ref-erence hospital. Material and methods: Type of study, retrospective evaluation of diagnostic tool, the size of the sample was calculated from the pilot simple and was 320 patients. We recolected 354 patients that urderwent both studies (CPRM and ERCP), to evaluate the correlation between both procedures we used Kappa variation. Results: 354 pacients were included, 226 women (63.8%), men 128 (36.2%), with mean age of 48 years. There was dilatation of the choledocus by CRMN in 60%, sensibility of 96, 7%, especificity of 40%, Kappa index 0,406 (IC 95%: 0, 32-0, 50) that indicates a moderate agreement. For the presence of choledocus calculi with CPRM was 54%, with sensibility of 72.6%, specificity of 68.3%, Kappa index 0.409 (IC 95%: 0.31-0.51), moderate agreement. For caculi in the gallbladder the kappa index was 0.246 weak. We identify 27% de false negatives for CRMN in detecting choledocusd lithiasis. Conclusions: The CPRM and ERCP had a moderate correlation according to the Kappa index detecting dilatation and choledocus calculi in our patients. The number of false negatives for cho-ledocolithiasis by CPRM leads us to seek in other prospective aleatory studies like endoscopic biliiopancreatic ultrasonopgraphy to compare the CPRM in patients with intermediate probability for choledoco-lithiasis

Prognostic value of the 6-gene OncoMasTR test in hormone receptor–positive HER2-negative early-stage breast cancer: Comparative analysis with standard clinicopathological factors

Aim: The aim of the study was to assess the prognostic performance of a 6-gene molecular score (OncoMasTR Molecular Score [OMm]) and a composite risk score (OncoMasTR Risk Score [OM]) and to conduct a within-patient comparison against four routinely used molecular and clinicopathological risk assessment tools: Oncotype DX Recurrence Score, Ki67, Nottingham Prognostic Index and Clinical Risk Category, based on the modified Adjuvant! Online definition and three risk factors: patient age, tumour size and grade. Methods: Biospecimens and clinicopathological information for 404 Irish women also previously enrolled in the Trial Assigning Individualized Options for Treatment [Rx] were provided by 11 participating hospitals, as the primary objective of an independent translational study. Gene expression measured via RT-qPCR was used to calculate OMm and OM. The prognostic value for distant recurrence-free survival (DRFS) and invasive disease-free survival (IDFS) was assessed using Cox proportional hazards models and Kaplan-Meier analysis. All statistical tests were two-sided ones. Results: OMm and OM (both with likelihood ratio statistic [LRS] P Discussion: Both OncoMasTR scores were significantly prognostic for DRFS and IDFS and provided additional prognostic information to the molecular and clinicopathological risk factors/tools assessed. OM was also the most accurate risk classification tool for identifying DR. A concise 6-gene signature with superior risk stratification was shown to increase prognosis reliability, which may help clinicians optimise treatment decisions. Trial registration: ClinicalTrials.gov NCT02050750 NCT00310180.</p