Molecular cloning and characterization of a thermostable esterase/lipase produced by a novel Anoxybacillus flavithermus strain

A thermophilic strain producing an extracellular esterase/lipase was isolated from a hot spring in Tǎşnad, Romania, and was identified phenotypically and by 16S rDNA sequencing as Anoxybacillus flavithermus (GenBank ID: JQ267733). The gene encoding the putative carboxyl esterase (GenBank ID: JX494348) was cloned by direct PCR amplification from genomic DNA. The protein, consisting of 246 amino acids and having a predicted molecular weight of 28.03 kDa, is encoded by an ORF of 741 bps. Expression was achieved in Escherichia coli and a recombinant protein with esterolytic activity and estimated molecular weight of 25 kDa was recovered and purified from the periplasmic fraction by IMAC. The purified enzyme, most active at 60-65°C and in the near-neutral range (pH 6.5-8), displayed a half-life at 60°C of about 5 h. Est/Lip displayed a relative tolerance to methanol, DMSO, acetonitrile, and low detergent concentrations (SDS, Triton) increased its thermostability. Highest activity was attained with p-nitrophenyl butyrate, but the enzyme was also able to hydrolyze long chain fatty acid esters, as well as triolein. The primary sequence and predicted tridimensional structure of the enzyme are very similar to those of other Anoxybacillus and Geobacillus carboxyl esterases in a distinct, recently described lipase family. Est/Lip was highly enantioselective, with preference for the (S)-enantiomer of substrates

Structural identifiability of dynamic systems biology models

22 páginas, 5 figuras, 2 tablas.-- This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.A powerful way of gaining insight into biological systems is by creating a nonlinear differential equation model, which usually contains many unknown parameters. Such a model is called structurally identifiable if it is possible to determine the values of its parameters from measurements of the model outputs. Structural identifiability is a prerequisite for parameter estimation, and should be assessed before exploiting a model. However, this analysis is seldom performed due to the high computational cost involved in the necessary symbolic calculations, which quickly becomes prohibitive as the problem size increases. In this paper we show how to analyse the structural identifiability of a very general class of nonlinear models by extending methods originally developed for studying observability. We present results about models whose identifiability had not been previously determined, report unidentifiabilities that had not been found before, and show how to modify those unidentifiable models to make them identifiable. This method helps prevent problems caused by lack of identifiability analysis, which can compromise the success of tasks such as experiment design, parameter estimation, and model-based optimization. The procedure is called STRIKE-GOLDD (STRuctural Identifiability taKen as Extended-Generalized Observability with Lie Derivatives and Decomposition), and it is implemented in a MATLAB toolbox which is available as open source software. The broad applicability of this approach facilitates the analysis of the increasingly complex models used in systems biology and other areasAFV acknowledges funding from the Galician government (Xunta de Galiza, Consellería de Cultura, Educación e Ordenación Universitaria http://www.edu.xunta.es/portal/taxonomy/term/206) through the I2C postdoctoral program, fellowship ED481B2014/133-0. AB and AFV were partially supported by grant DPI2013-47100-C2-2-P from the Spanish Ministry of Economy and Competitiveness (MINECO). AFV acknowledges additional funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement No 686282 (CanPathPro). AP was partially supported through EPSRC projects EP/M002454/1 and EP/J012041/1.Peer reviewe

TDCS calculations for the ionization of H

Triple differential cross sections (TDCSs) are calculated for the ionization of the hydrogen molecule by electron impact with asymmetric and symmetric kinematical configuration, respectively in coplanar arrangement and in the plane perpendicular to the incident direction. In the scattering plane we explore the angular distribution of the TDCS as well as the dependence of the cross section on the internuclear separation. In the perpendicular plane the TDCSs are investigated as a function of the angular separation between the outgoing electrons and for an equal energy sharing between them

Electron impact ionization of diatomic molecules

Cross sections for the ionization of N2, CO and O2 diatomic molecules by electron impact are calculated. The applied distorted wave model is based on our previous studies for positron impact, the molecular orbitals being described by Gaussian wavefunctions. Our study emphasizes the importance of electron exchange and of using correct distorted waves for the ejected electron

Visualization of formal specifications for understanding and debugging an industrial DSL

In this work we report on our proof of concept of a generic approach: visualized formal specification of a Domain Specific Language (DSL) can be used for debugging, understanding, and impact analysis of the DSL programs. In our case study we provide a domain-specific visualization for the Event-B specification of a real-life industrial DSL and perform a user study among DSL engineers to discover opportunities for its application. In this paper, we explain the rationale behind our visualization design, discuss the technical challenges of its realization and how these challenges were solved using the Model Driven Engineering (MDE) techniques. Based on the positive feedback of the user study, we present our vision on how this successful experience can be reused and the approach can be generalized for other DSLs

On the Dichotomy of Debugging Behavior Among Programmers

Debugging is an inevitable activity in most software projects, often difficult and more time-consuming than expected, giving it the nickname the “dirty little secret of computer science.” Surprisingly, we have little knowledge on how software engineers debug software problems in the real world, whether they use dedicated debugging tools, and how knowledgeable they are about debugging. This study aims to shed light on these aspects by following a mixed-methods research approach. We conduct an online survey capturing how 176 developers reflect on debugging. We augment this subjective survey data with objective observations on how 458 developers use the debugger included in their integrated development environments (IDEs) by instrumenting the popular ECLIPSE and INTELLIJ IDEs with the purpose-built plugin WATCHDOG 2.0. To clarify the insights and discrepancies observed in the previous steps, we followed up by conducting interviews with debugging experts and regular debugging users. Our results indicate that IDE-provided debuggers are not used as often as expected, because “printf debugging” remains a feasible choice for many programmers. Furthermore, both knowledge and use of advanced debugging features are low. Our results call for strengthening hands-on debugging experience in computer science curricula and have already refined the implementation of modern IDE debuggers.Green Open Access added to TU Delft Institutional Repository ‘You share, we take care!’ – Taverne project https://www.openaccess.nl/en/you-share-we-take-care Otherwise as indicated in the copyright section: the publisher is the copyright holder of this work and the author uses the Dutch legislation to make this work public.Software Engineerin