Respiratory versatility in Desulfovibrio desulfuricans ATCC 27774 – a proteomic approach

Poster presented at the Bacterial Electron Transfer Processes and their Regulation Meeting, European Federation of Biotechnology Microbial Physiology Section, 15-18 March 2015, Vimeiro, Portugal

Preparation of biodegradable materials by reactive extrusion

This work aimed to prepare biodegradable polymeric materials based on blends of a synthetic high density polyethylene (HDPE) and biodegradable polymers such as polylactic acid (PCL) and poly(caprolactone) (PLA), in a co-rotating twin-screw extruder. A polyethylene modified with maleic anhydride was used as compatibiliser. The mechanical results showed that the addition of PLA improves the blends stiffness while the addition of PCL leads to materials with a greater elongation at break and a lower Young modulus. This feature is related with the mechanical properties of each material as well as the adhesion between them. Concerning the biodegradability tests, it was found that HDPE/PCL blend presents the highest degree of biodegradability

A genetic code alteration generates a proteome of high diversity in the human pathogen Candida albicans

Background - Genetic code alterations have been reported in mitochondrial, prokaryotic, and eukaryotic cytoplasmic translation systems, but their evolution and how organisms cope and survive such dramatic genetic events are not understood. Results - Here we used an unusual decoding of leucine CUG codons as serine in the main human fungal pathogen Candida albicans to elucidate the global impact of genetic code alterations on the proteome. We show that C. albicans decodes CUG codons ambiguously and tolerates partial reversion of their identity from serine back to leucine on a genome-wide scale. Conclusion - Such codon ambiguity expands the proteome of this human pathogen exponentially and is used to generate important phenotypic diversity. This study highlights novel features of C. albicans biology and unanticipated roles for codon ambiguity in the evolution of the genetic code.publishe

A Genetic Code Alteration Is a Phenotype Diversity Generator in the Human Pathogen Candida albicans

BACKGROUND: The discovery of genetic code alterations and expansions in both prokaryotes and eukaryotes abolished the hypothesis of a frozen and universal genetic code and exposed unanticipated flexibility in codon and amino acid assignments. It is now clear that codon identity alterations involve sense and non-sense codons and can occur in organisms with complex genomes and proteomes. However, the biological functions, the molecular mechanisms of evolution and the diversity of genetic code alterations remain largely unknown. In various species of the genus Candida, the leucine CUG codon is decoded as serine by a unique serine tRNA that contains a leucine 5'-CAG-3'anticodon (tRNA(CAG)(Ser)). We are using this codon identity redefinition as a model system to elucidate the evolution of genetic code alterations. METHODOLOGY/PRINCIPAL FINDINGS: We have reconstructed the early stages of the Candida genetic code alteration by engineering tRNAs that partially reverted the identity of serine CUG codons back to their standard leucine meaning. Such genetic code manipulation had profound cellular consequences as it exposed important morphological variation, altered gene expression, re-arranged the karyotype, increased cell-cell adhesion and secretion of hydrolytic enzymes. CONCLUSION/SIGNIFICANCE: Our study provides the first experimental evidence for an important role of genetic code alterations as generators of phenotypic diversity of high selective potential and supports the hypothesis that they speed up evolution of new phenotypes

Penicillium crustosum as a potential OTA producer - new insights from whole - genome sequencing of strain MUM 16.125

Ochratoxin A (OTA) is a well-studied mycotoxin that poses severe health risks. OTA is mainly produced by Aspergillus and Penicillium species associated with food spoilage and it is present in a wide diversity of food and feed products. Recent studies have reported the presence of OTA in food matrices where known OTA producers are not present1,2. For that reason, other species such as P. crustosum are now being considered. A recent study using comparative genomic analysis3 clarified the OTA biosynthetic gene cluster composition. In order to gain insight into the secondary metabolism of P. crustosum, this study aimed to sequence and explore the complete genome of strain MUM 16.125. This strain was isolated from cheese rind sample contaminated with OTA in which no known OTA producers were present1. The genome assembly comprises 199 contigs with a total length of 30.95 Mb and contains 10975 predicted protein-coding genes. In total, 109 gene clusters potentially related with secondary metabolism were identified, including putative gene clusters for penitrem, clavaric acid or naphthopyrones biosynthesis. Nevertheless, no evidence of an OTA biosynthetic gene cluster was found. A total of 83 complete and 49 partial protein sequences from published OTA biosynthetic genes from 11 Aspergillus and 3 Penicillium species were queried against the predicted P. crustosum proteins. Only 3 strong matches were found (to a short partial P. verrucosum PKS and 2 P. thymicola chloroperoxidases) but matches to complete key genes were absent. Considering these findings, it appears that strain MUM 16.125 lacks the most common genetic pathway to produce OTA, providing important information relevant to understand the role of P. crustosum as putative OTA producer. Nevertheless, the additional secondary metabolism gene clusters found (such as penitrem, clavaric acid or naphthopyrones) highlight the potential of this strain for metabolite production, including other mycotoxins or compounds with antioxidant, anticancer or antibiotic properties.This study was supported by the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of CEB (UID/BIO/04469/2019) and iBiMED (UIDB/04501/2020) units; and by CANCYL (POCI-01-0145-FEDER-031849) and GenomePT (POCI-01-0145-FEDER-022184) projectsinfo:eu-repo/semantics/publishedVersio

Codon-triplet context unveils unique features of the Candida albicans protein coding genome

<p>Abstract</p> <p>Background</p> <p>The evolutionary forces that determine the arrangement of synonymous codons within open reading frames and fine tune mRNA translation efficiency are not yet understood. In order to tackle this question we have carried out a large scale study of codon-triplet contexts in 11 fungal species to unravel associations or relationships between codons present at the ribosome A-, P- and E-sites during each decoding cycle.</p> <p>Results</p> <p>Our analysis unveiled high bias within the context of codon-triplets, in particular strong preference for triplets of identical codons. We have also identified a surprisingly large number of codon-triplet combinations that vanished from fungal ORFeomes. <it>Candida albicans </it>exacerbated these features, showed an unbalanced tRNA population for decoding its pool of codons and used near-cognate decoding for a large set of codons, suggesting that unique evolutionary forces shaped the evolution of its ORFeome.</p> <p>Conclusion</p> <p>We have developed bioinformatics tools for large-scale analysis of codon-triplet contexts. These algorithms identified codon-triplets context biases, allowed for large scale comparative codon-triplet analysis, and identified rules governing codon-triplet context. They could also detect alterations to the standard genetic code.</p

Species-Specific Codon Context Rules Unveil Non-Neutrality Effects of Synonymous Mutations

Codon pair usage (codon context) is a species specific gene primary structure feature whose evolutionary and functional roles are poorly understood. The data available show that codon-context has direct impact on both translation accuracy and efficiency, but one does not yet understand how it affects these two translation variables or whether context biases shape gene evolution.Here we study codon-context biases using a set of 72 orthologous highly conserved genes from bacteria, archaea, fungi and high eukaryotes to identify 7 distinct groups of codon context rules. We show that synonymous mutations, i.e., neutral mutations that occur in synonymous codons of codon-pairs, are selected to maintain context biases and that non-synonymous mutations, i.e., non-neutral mutations that alter protein amino acid sequences, are also under selective pressure to preserve codon-context biases.Since in vivo studies provide evidence for a role of codon context on decoding fidelity in E. coli and for decoding efficiency in mammalian cells, our data support the hypothesis that, like codon usage, codon context modulates the evolution of gene primary structure and fine tunes the structure of open reading frames for high genome translational fidelity and efficiency in the 3 domains of life

Low level genome mistranslations deregulate the transcriptome and translatome and generate proteotoxic stress in yeast

Organisms use highly accurate molecular processes to transcribe their genes and a variety of mRNA quality control and ribosome proofreading mechanisms to maintain intact the fidelity of genetic information flow. Despite this, low level gene translational errors induced by mutations and environmental factors cause neurodegeneration and premature death in mice and mitochondrial disorders in humans. Paradoxically, such errors can generate advantageous phenotypic diversity in fungi and bacteria through poorly understood molecular processes.publishe

Relação familiar em diálogo com o desenvolvimento psíquico da criança: Análise sobre a visão de Coraline

A análise foi construída pensando na influência das relações familiares sobre as crianças, utilizando-se das categorias fuga e frustração procurou adentrar ao aparelho psíquico das crianças. Para isso, a revisão de literatura juntamente com observação sobre o filme Coraline e o Mundo Secreto de Henry Selick foram utilizados como método. Analisando a construção do aparelho psíquico da personagem principal em diálogo com as categorias de comportamento selecionadas e tendo em vista a relevância do tema, ao entender que todas as crianças passam pelo processo de dialogar suas vontades com as demandas externas. Durante a análise, concluiu-se, através da relação de Coraline com seus pais, que a falta de atenção vinda dos pais pode exacerbar as frustrações presentes no contato da criança com as normas sociais e conduzir para uma fuga a um mundo imaginário mais satisfatório. Assim, de maneira alternativa, foi contemplada a possibilidade de aplicar a parentalidade positiva como forma de produzir um desenvolvimento saudável à criança.  Foi também pontuada potencialidades advindas das castrações por meio da negativa a pedidos exacerbados, a qual pode promover maior sucesso na interação do mundo interior com o exterior, na fase adulta.Palavras-chaves: família; fuga; frustração; parentalidade; aparelho psíquico.

Construction of effective disposable biosensors for point of care testing of nitrite

© 2015. This manuscript version is made available under the CC-BY-NC-ND 4.0 license http://creativecommons.org/licenses/by-nc-nd/4.0/"In this paper we aim to demonstrate, as a proof-of-concept, the feasibility of the mass production of effective point of care tests for nitrite quantification in environmental, food and clinical samples. Following our previous work on the development of third generation electrochemical biosensors based on the ammonia forming nitrite reductase (ccNiR), herein we reduced the size of the electrodes’ system to a miniaturized format, solved the problem of oxygen interference and performed simple quantification assays in real samples. In particular, carbon paste screen printed electrodes (SPE) were coated with a ccNiR/carbon ink composite homogenized in organic solvents and cured at low temperatures. The biocompatibility of these chemical and thermal treatments was evaluated by cyclic voltammetry showing that the catalytic performance was higher with the combination acetone and a 40 °C curing temperature. The successful incorporation of the protein in the carbon ink/solvent composite, while remaining catalytically competent, attests for ccNiR’s robustness and suitability for application in screen printed based biosensors. Because the direct electrochemical reduction of molecular oxygen occurs when electroanalytical measurements are performed at the negative potentials required to activate ccNiR (ca. -0.4 V vs Ag/AgCl), an oxygen scavenging system based on the coupling of glucose oxidase and catalase activities was successfully used. This enabled the quantification of nitrite in different samples (milk, water, plasma and urine) in a straightforward way and with small error (1 – 6%). The sensitivity of the biosensor towards nitrite reduction under optimized conditions was 0.55 A M-1 cm-2 with a linear response range 0.7 – 370 μM.