Insights into human carboxylesterase 2 stability and activity in vivo and in vitro

Dissertation to obtain a Master Degree in Biotechnology

Structural stability of adenylate kinase from the sulfate-reducing bacteria Desulfovibrio gigas

Biophysical Chemistry 110 (2004) 83–92

Automatic learning for the classification of chemical reactions and in statistical thermodynamics

This Thesis describes the application of automatic learning methods for a) the classification of organic and metabolic reactions, and b) the mapping of Potential Energy Surfaces(PES). The classification of reactions was approached with two distinct methodologies: a representation of chemical reactions based on NMR data, and a representation of chemical reactions from the reaction equation based on the physico-chemical and topological features of chemical bonds. NMR-based classification of photochemical and enzymatic reactions. Photochemical and metabolic reactions were classified by Kohonen Self-Organizing Maps (Kohonen SOMs) and Random Forests (RFs) taking as input the difference between the 1H NMR spectra of the products and the reactants. The development of such a representation can be applied in automatic analysis of changes in the 1H NMR spectrum of a mixture and their interpretation in terms of the chemical reactions taking place. Examples of possible applications are the monitoring of reaction processes, evaluation of the stability of chemicals, or even the interpretation of metabonomic data. A Kohonen SOM trained with a data set of metabolic reactions catalysed by transferases was able to correctly classify 75% of an independent test set in terms of the EC number subclass. Random Forests improved the correct predictions to 79%. With photochemical reactions classified into 7 groups, an independent test set was classified with 86-93% accuracy. The data set of photochemical reactions was also used to simulate mixtures with two reactions occurring simultaneously. Kohonen SOMs and Feed-Forward Neural Networks (FFNNs) were trained to classify the reactions occurring in a mixture based on the 1H NMR spectra of the products and reactants. Kohonen SOMs allowed the correct assignment of 53-63% of the mixtures (in a test set). Counter-Propagation Neural Networks (CPNNs) gave origin to similar results. The use of supervised learning techniques allowed an improvement in the results. They were improved to 77% of correct assignments when an ensemble of ten FFNNs were used and to 80% when Random Forests were used. This study was performed with NMR data simulated from the molecular structure by the SPINUS program. In the design of one test set, simulated data was combined with experimental data. The results support the proposal of linking databases of chemical reactions to experimental or simulated NMR data for automatic classification of reactions and mixtures of reactions. Genome-scale classification of enzymatic reactions from their reaction equation. The MOLMAP descriptor relies on a Kohonen SOM that defines types of bonds on the basis of their physico-chemical and topological properties. The MOLMAP descriptor of a molecule represents the types of bonds available in that molecule. The MOLMAP descriptor of a reaction is defined as the difference between the MOLMAPs of the products and the reactants, and numerically encodes the pattern of bonds that are broken, changed, and made during a chemical reaction. The automatic perception of chemical similarities between metabolic reactions is required for a variety of applications ranging from the computer validation of classification systems, genome-scale reconstruction (or comparison) of metabolic pathways, to the classification of enzymatic mechanisms. Catalytic functions of proteins are generally described by the EC numbers that are simultaneously employed as identifiers of reactions, enzymes, and enzyme genes, thus linking metabolic and genomic information. Different methods should be available to automatically compare metabolic reactions and for the automatic assignment of EC numbers to reactions still not officially classified. In this study, the genome-scale data set of enzymatic reactions available in the KEGG database was encoded by the MOLMAP descriptors, and was submitted to Kohonen SOMs to compare the resulting map with the official EC number classification, to explore the possibility of predicting EC numbers from the reaction equation, and to assess the internal consistency of the EC classification at the class level. A general agreement with the EC classification was observed, i.e. a relationship between the similarity of MOLMAPs and the similarity of EC numbers. At the same time, MOLMAPs were able to discriminate between EC sub-subclasses. EC numbers could be assigned at the class, subclass, and sub-subclass levels with accuracies up to 92%, 80%, and 70% for independent test sets. The correspondence between chemical similarity of metabolic reactions and their MOLMAP descriptors was applied to the identification of a number of reactions mapped into the same neuron but belonging to different EC classes, which demonstrated the ability of the MOLMAP/SOM approach to verify the internal consistency of classifications in databases of metabolic reactions. RFs were also used to assign the four levels of the EC hierarchy from the reaction equation. EC numbers were correctly assigned in 95%, 90%, 85% and 86% of the cases (for independent test sets) at the class, subclass, sub-subclass and full EC number level,respectively. Experiments for the classification of reactions from the main reactants and products were performed with RFs - EC numbers were assigned at the class, subclass and sub-subclass level with accuracies of 78%, 74% and 63%, respectively. In the course of the experiments with metabolic reactions we suggested that the MOLMAP / SOM concept could be extended to the representation of other levels of metabolic information such as metabolic pathways. Following the MOLMAP idea, the pattern of neurons activated by the reactions of a metabolic pathway is a representation of the reactions involved in that pathway - a descriptor of the metabolic pathway. This reasoning enabled the comparison of different pathways, the automatic classification of pathways, and a classification of organisms based on their biochemical machinery. The three levels of classification (from bonds to metabolic pathways) allowed to map and perceive chemical similarities between metabolic pathways even for pathways of different types of metabolism and pathways that do not share similarities in terms of EC numbers. Mapping of PES by neural networks (NNs). In a first series of experiments, ensembles of Feed-Forward NNs (EnsFFNNs) and Associative Neural Networks (ASNNs) were trained to reproduce PES represented by the Lennard-Jones (LJ) analytical potential function. The accuracy of the method was assessed by comparing the results of molecular dynamics simulations (thermal, structural, and dynamic properties) obtained from the NNs-PES and from the LJ function. The results indicated that for LJ-type potentials, NNs can be trained to generate accurate PES to be used in molecular simulations. EnsFFNNs and ASNNs gave better results than single FFNNs. A remarkable ability of the NNs models to interpolate between distant curves and accurately reproduce potentials to be used in molecular simulations is shown. The purpose of the first study was to systematically analyse the accuracy of different NNs. Our main motivation, however, is reflected in the next study: the mapping of multidimensional PES by NNs to simulate, by Molecular Dynamics or Monte Carlo, the adsorption and self-assembly of solvated organic molecules on noble-metal electrodes. Indeed, for such complex and heterogeneous systems the development of suitable analytical functions that fit quantum mechanical interaction energies is a non-trivial or even impossible task. The data consisted of energy values, from Density Functional Theory (DFT) calculations, at different distances, for several molecular orientations and three electrode adsorption sites. The results indicate that NNs require a data set large enough to cover well the diversity of possible interaction sites, distances, and orientations. NNs trained with such data sets can perform equally well or even better than analytical functions. Therefore, they can be used in molecular simulations, particularly for the ethanol/Au (111) interface which is the case studied in the present Thesis. Once properly trained, the networks are able to produce, as output, any required number of energy points for accurate interpolations.

Evaluation of the potential of translocated common cockle for ecological risk assessment studies: bioaccumulation and biomarkers test

Thesis submitted to the Faculdade de Ciências e Tecnologia to obtain the Master’s degree in Environmental Engineering, profile in Ecological Engineering

Gammaretroviral and lentiviral vectors for gene therapy: stability and inactivation mechanisms

Dissertation presented to obtain a Ph.D degree in Engineering and Technology Sciences, Gene Therapy at the Instituto de Tecnologia Quimica e Biológica, Universidade Nova de Lisboa

A proteomic approach toward the selection of proteins with enhanced intrinsic conformational stability

Journal of Proteome Research (2006)5: 2720-2726

Metal ions modulate the folding and stability of the tumor suppressor protein S100A2

Febs Journal (2009)276:1776-1786

In vitro studies of gum arabic-coated magnetic nanoparticles with mammalian cell cultures

Dissertação apresentada na Faculdade de Ciências e Tecnologia da Universidade Nova de Lisboa para obtenção do grau de Mestre em Biotecnologia

Relação entre a catastrofização da dor, percepção da intensidade da dor e incapacidade funcional em utentes com cdor crónica cervical

RESUMO: Introdução/Objectivo: A influência dos factores psicossociais, e nomeadamente da catastrofização da dor, na percepção da intensidade da dor e na incapacidade funcional, auto-reportada por indivíduos com dor crónica cervical (DCC), tem sido alvo de estudo em vários países, evidenciando o constructo multidimensional da DCC. Neste sentido, esta investigação pretende estudar se a catastrofização da dor, é uma variável preditiva relativamente à percepção da intensidade da dor, e à incapacidade funcional. Secundariamente procurou-se averiguar se as relações encontradas se mantêm estáveis antes e após exposição a uma intervenção em fisioterapia. Metodologia: Neste estudo participaram 40 indivíduos com DCC de origem músculo-esquelética e causa não traumática, que foram expostos a uma intervenção em fisioterapia no Centro de Medicina de Reabilitação do Alcoitão e na Clinica AlcaisFisio, que cumpriram os critérios de inclusão e aceitarem participar livremente no mesmo. A recolha de dados realizou-se em dois momentos distintos, antes e após exposição à intervenção em fisioterapia. A catastrofização da dor foi avaliada por meio da Escala de Catastrofização da Dor (PCS), a intensidade da dor pela Escala Numérica da Dor (END), sendo realizada a medição da incapacidade funcional através do Neck Disability Index versão Portuguesa (NDI-PT). A análise estatística incluiu duas fases: fase descritiva e fase inferencial. Foram desenvolvidos modelos de regressão linear com vista a testar o poder preditivo da catastrofização da dor sobre a intensidade da dor e a incapacidade funcional. O nível de significância para o qual os valores se consideraram satisfatórios foi de p<0,05. O tratamento dos dados foi realizado no software PASW versão 18. Resultados: Observou-se que existe uma relação moderada, positiva e significativa, nos dois momentos de avaliação, entre a catastrofização da dor e a percepção da intensidade da dor (p<0,001), apresentando um poder preditivo de 27,9% e 46,7%, das pontuações da intensidade da dor, antes e após exposição à intervenção em fisioterapia, espectivamente. Observou-se que a catastrofização da dor tem uma relação forte, positiva e significativa com a incapacidade funcional, nos dois momentos de avaliação (p<0,001), predizendo 51,8% e 61,8%, das pontuações da incapacidade funcional, antes e após exposição à intervenção em fisioterapia, respectivamente. Conclusão: A catastrofização da dor é um factor psicossocial que apresenta relação moderada com a percepção da intensidade da dor, e forte com a incapacidade funcional auto-reportada por indivíduos com DCC de origem músculo-esquelética e causa não traumática, antes e após exposição à intervenção em fisioterapia. Os resultados do estudo sugerem, assim, uma importante influência da catastrofização da dor sobre a percepção da intensidade da dor e a incapacidade funcional em indivíduos com DCC, realçando o constructo multidimensional da DCC. ------------ABSTRACT: Background and Purpose: The influence of psychosocial factors, particularly, the pain catastrophizing, on pain intensity and functional disability in individuals with chronic neck pain (CNP) has been report among recent research literature. The first aim of this research was to verify the predictive value of pain catastrophizing on pain intensity and patient’s functional disability. Secondly it aimed to verify the stability of these relations before and after a physiotherapy treatment. Methodology: A sample of 40 subjects with CNP of musculoskeletal and non-traumatic causes was recruited from the patient’s list of two private clinics in Lisbon district following verification of the inclusion criteria. All participants agree to participate in the study and signed a consent form. Data was collected immediately before and after a period of physiotherapy treatment. Pain catastrophizing was assessed by the Pain Catastrophizing Scale (PCS-PT), patient perception of pain intensity was measured by the Numerical Rating Scale (NRS), and functional disability was measured through the Neck Disability Index (NDI-PT). Data was analyzed through descriptive and inferential statistics. Linear regression models were developed in order to test the predictive power of pain catastrophizing on pain intensity and functional disability. The minimal level of significance established was p<0,05. Data analysis was performed using the software PASW version 18. Results: A positive moderate relationship between pain catastrophizing and pain intensity was founded in both moments, before and after physiotherapy intervention, of data collection (p<0,001) with a predictive power of 27,9% and 46,7%, respectively. A positive strong relationship between pain catastrophizing and functional disability was founded in both moments, before and after physiotherapy intervention, of data collection (p<0,001), with a predictive power of 51,8% and 61,8%, respectively. Conclusion: Pain catastrophizing is a psychosocial factor that is correlated moderately with the perception of pain intensity and strongly with self-reported functionaldisability for individuals with CNP musculoskeletal origin and non-traumatic causes,before and after a physiotherapy intervention. The results of this study suggest that pain catastrophizing has an important influence on the report levels of pain intensity and functional disability in CNP patients. These results also emphasize the multidimensional nature of chronic neck pain.

A score test for non-nested hypotheses with applications to discrete data models

This paper suggests that a convenient score test against non-nested alternatives can be constructed from the linear combination of the likelihood functions of the competing models. It is shown that this procedure is essentially a test for the correct specification of the conditional distribution of the variable of interest.

Using gold nanoparticles in protein crystallography: studies in crystal growth and derivatization

Dissertation for the Master Degree in Structural and Functional Biochemistry

Insight into the multicopper oxidases stability

Dissertation presented to obtain the PhD degree in Biochemistry

Weekly tracking of stability of the flow of conversations into the subprocesses of the Last Planner System

21st Annual Conference of the International Group for Lean Construction – IGLC 21 – Fortaleza, Brazil

Protein stability in a proteomic perspective

Dissertation presented to obtain the Ph.D degree in Biochemistry

O biofeedback cinemático na realização de exercícios escápulo-torácicos

RESUMO: O Biofeedback cinemático na realização de exercícios escápulo-torácicos Patrícia Ribeiro; Ricardo Matias Desenho do estudo: Estudo quantitativo, experimental, prospetivo de fator único. Objetivos: Determinar a efetividade do biofeedback cinemático tridimensional (3D) em tempo real, como informação de retorno extrínseca, no reconhecimento da zona neutra da omoplata, em indivíduos saudáveis; medir o desempenho temporal, através do tempo que os sujeitos demoram a atingir a posição pretendida, e o desempenho no percurso, através da diferença entre o percurso efetuado e aquele que seria o percurso mais curto, sem Informação de Retorno Extrínseca (IRE), com Informação de Retorno Extrínseca clinica verbal e palpatória (IRE-VP) e com informação de retorno extrínseca visual cinemática (IRE-VC). Enquadramento: Reconhecer aquela que deve ser a posição adequada da omoplata e tomar consciência da importância dessa posição para a estabilidade e correto funcionamento da articulação gleno-umeral é uma tarefa difícil, pois a grande maioria do indivíduos não têm os conhecimentos necessários. É reconhecido o contributo que a IRE tem na reaprendizagem motora, em especial na primeira fase da aprendizagem motora – a consciencialização. Vários estudos se têm debruçado em avaliar a influencia da IRE na reabilitação de indivíduos com disfunção do ombro, tendo-se limitado ao formato electromiográfico. No sentido de encontrar novas estratégias para facilitar o processo de reaprendizagem motora nestes indivíduos, foi escolhido biofeedback cinemático tridimensional. Métodos: Participaram no estudo 60 sujeitos, sem queixas de dor no ombro, com idades compreendidas entre os 18 e os 54 anos, com um valor médio de 21,66 anos (±5.89), divididos aleatoriamente em três grupos com uma tarefa comum: o reconhecimento da zona neutra da omoplata. Um grupo não recebeu qualquer tipo de IRE - Grupo Controlo (GC), outro grupo recebeu IRE-VP e outro recebeu IRE-VC. Para efeitos de análise da performance foram consideradas duas variáveis: tempo e qualidade da execução. A abordagem estatística destas variáveis consistiu numa análise de variância simples e num teste de comparações múltiplas. Resultados: Observámos que o grupo controlo e o grupo IRE-VC efetuaram a tarefa pedida em menos tempo que o grupo IRE-VP (p<0.01) e que o grupo IRE-VC obteve melhores resultados no que respeita à qualidade da execução da tarefa do que o GC e o grupo IRE-VP (p<0.01). Conclusão: Pelos resultados obtidos, na tarefa de reconhecimento da zona neutra da omoplata, podemos concluir que o biofeedback cinemático é mais efetivo na facilitação da obtenção de melhor qualidade de execução da tarefa do que o feedback clinico ou a inexistência de feedback e igualmente efetivo à inexistência de IRE no que respeita ao tempo demorado a reconhecer essa posição. -------------- ABSTRACT:Study Design: Single factor experimental design. Objectives: To determine the effectiveness of real time cinematic biofeedback as extrinsic information in the recognition of the neutral zone of the scapula in healthy subjects; to measure the temporal performance and quality of the execution without extrinsic feedback, with clinical feedback and with cinematic feedback. Background: To recognize the proper scapula position and to be aware of the importance of this position to the stability and correct function of the gleno-umeral joint is a difficult task, because most people don’t have the necessary knowledge. It is known that extrinsic feedback is effective in motor learning, specially in the early stages. There are several studies that evaluated the influence of extrinsic feedback in rehabilitation of shoulder dysfunctions, but only in the electromyography format. Material and Methods: Sixty healthy subjects were randomly divided into 3 groups that received no feedback (group 1 – control group), verbal and palpatory feedback (group 2) and real time cinematic feedback (group 3). The performance variables considered – time and quality of execution - were statistically analyzed with ANOVA one-way and multiple comparisons test. Results: Control group and group 3 executed the task in less time that group 2 (p<0.01) and group 3 executed the task with better quality than control group and group 2 (p<0.01). Conclusions: From the results of this study we conclude that in the studied task, the cinematic feedback is more effective to facilitate a better quality of execution than no feedback or clinical feedback and, regarding to time taken to attain the requested position, equally effective to no feedback.