Relevance of arginine residues in Cu(II)-induced DNA breakage and Proteinase K resistance of H1 histones

This work analyzes the involvement of arginines in copper/H2O2-induced DNA breakage. Copper is a highly redox active metal which has been demonstrated to form compounds with arginines. For this aim we used mixtures of pGEM3 DNA plasmid and two types of H1 histones which differ only in their arginine content. The sperm H1 histone from the annelid worm Chaetopterus variopedatus (arginine content 12.6 mol% K/R ratio 2) and the somatic H1 histone from calf thymus (arginine content 1.8 mol% and K/R ratio 15). Copper/H2O2-induced DNA breakage was observed only in presence of sperm H1 histones, but it was more relevant for the native molecule than for the deguanidinated derivative (K/R ratio 14), in which 80% of arginine residues were converted to ornithine. Further, copper induced proteinase K resistance and increase of DNA binding affinity on native sperm H1 histones. These observations are consistent with a copper induced reorganization of the side-chains of arginine residues. Copper, instead, did not affect DNA binding affinity of somatic and deguanidinated H1 histones, which show similar K/R ratio and DNA binding mode. These results indicate that arginine residues could affect these H1 histones properties and provide new insights into copper toxicity mechanisms

SDS-resistant Active and Thermostable Dimers Are Obtained from the Dissociation of Homotetrameric β-Glycosidase from Hyperthermophilic Sulfolobus solfataricus in SDS

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Redox stress proteins are involved in adaptation response of the hyperthermoacidophilic archaeon Sulfolobus solfataricus to nickel challenge

<p>Abstract</p> <p>Background</p> <p>Exposure to nickel (Ni) and its chemical derivatives has been associated with severe health effects in human. On the contrary, poor knowledge has been acquired on target physiological processes or molecular mechanisms of this metal in model organisms, including Bacteria and Archaea. In this study, we describe an analysis focused at identifying proteins involved in the recovery of the archaeon <it>Sulfolobus solfataricus </it>strain MT4 from Ni-induced stress.</p> <p>Results</p> <p>To this purpose, <it>Sulfolobus solfataricus </it>was grown in the presence of the highest nickel sulphate concentration still allowing cells to survive; crude extracts from treated and untreated cells were compared at the proteome level by using a bi-dimensional chromatography approach. We identified several proteins specifically repressed or induced as result of Ni treatment. Observed up-regulated proteins were largely endowed with the ability to trigger recovery from oxidative and osmotic stress in other biological systems. It is noteworthy that most of the proteins induced following Ni treatment perform similar functions and a few have eukaryal homologue counterparts.</p> <p>Conclusion</p> <p>These findings suggest a series of preferential gene expression pathways activated in adaptation response to metal challenge.</p

A Substrate-induced Switch in the Reaction Mechanism of a Thermophilic Esterase KINETIC EVIDENCES AND STRUCTURAL BASIS

The reaction mechanism of the esterase 2 (EST2) from Alicyclobacillus acidocaldarius was studied at the kinetic and structural level to shed light on the mechanism of activity and substrate specificity increase previously observed in its double mutant M211S/R215L. In particular, the values of kinetic constants (k1, k(-1), k2, and k3) along with activation energies (E1, E(-1), E2, and E3) were measured for wild type and mutant enzyme. The previously suggested substrate-induced switch in the reaction mechanism from kcat=k3 with a short acyl chain substrate (p-nitrophenyl hexanoate) to kcat=k2 with a long acyl chain substrate (p-nitrophenyl dodecanoate) was validated. The inhibition afforded by an irreversible inhibitor (1-hexadecanesulfonyl chloride), structurally related to p-nitrophenyl dodecanoate, was studied by kinetic analysis. Moreover the three-dimensional structure of the double mutant bound to this inhibitor was determined, providing essential information on the enzyme mechanism. In fact, structural analysis explained the observed substrate-induced switch because of an inversion in the binding mode of the long acyl chain derivatives with respect to the acyl- and alcohol-binding sites

New Cross-Talk Layer between Ultraconserved Non-Coding RNAs, MicroRNAs and Polycomb Protein YY1 in Bladder Cancer

MicroRNAs (miRNAs) are highly conserved elements in mammals, and exert key regulatory functions. Growing evidence shows that miRNAs can interact with another class of non-coding RNAs, so-called transcribed ultraconserved regions (T-UCRs), which take part in transcriptional, post-transcriptional and epigenetic regulation processes. We report here the interaction of miRNAs and T-UCRs as a network modulating the availability of these non-coding RNAs in bladder cancer cells. In our cell system, antagomiR-596 increased the expression of T-UCR 201+. Moreover, T-UCR 8+ silencing increased miR-596 expression, which in turn reduced total T-UCR 283+, showing that the perturbation of one element in this network changes the expression of other interactors. In addition, we identify the polycomb protein Yin Yang 1 (YY1) as mediator of binding between miR-596 and T-UCR 8+. These new findings describe for the first time a network between T-UCRs, miRNAs and YY1 protein, highlighting the existence of an additional layer of gene expression regulation

Long non-coding RNA containing ultraconserved genomic region 8 promotes bladder cancer tumorigenesis

Ultraconserved regions (UCRs) have been shown to originate non-coding RNA transcripts (T-UCRs) that have different expression profiles and play functional roles in the pathophysiology of multiple cancers. The relevance of these functions to the pathogenesis of bladder cancer (BlCa) is speculative. To elucidate this relevance, we first used genome-wide profiling to evaluate the expression of T-UCRs in BlCa tissues. Analysis of two datasets comprising normal bladder tissues and BlCa specimens with a custom T-UCR microarray identified ultraconserved RNA (uc.) 8+ as the most upregulated T-UCR in BlCa tissues, although its expression was lower than in pericancerous bladder tissues. These results were confirmed on BlCa tissues by real-time PCR and by in situ hybridization. Although uc.8+ is located within intron 1 of CASZ1, a zinc-finger transcription factor, the transcribed non-coding RNA encoding uc.8+ is expressed independently of CASZ1. In vitro experiments evaluating the effects of uc.8+ silencing, showed significantly decreased capacities for cancer cell invasion, migration, and proliferation. From this, we proposed and validated a model of interaction in which uc.8+ shuttles from the nucleus to the cytoplasm of BlCa cells, interacts with microRNA (miR)-596, and cooperates in the promotion and development of BlCa. Using computational analysis, we investigated the miR-binding domain accessibility, as determined by base-pairing interactions within the uc.8+ predicted secondary structure, RNA binding affinity, and RNA species abundance in bladder tissues and showed that uc.8+ is a natural decoy for miR-596. Thus uc.8+ upregulation results in increased expression of MMP9, increasing the invasive potential of BlCa cells. These interactions between evolutionarily conserved regions of DNA suggest that natural selection has preserved this potentially regulatory layer that uses RNA to modulate miR levels, opening up the possibility for development of useful markers for early diagnosis and prognosis as well as for development of new RNA-based cancer therapies

Risk assessment of honey bee stressors based on in silico analysis of molecular interactions

Abstract A global decline of the honey bee Apis mellifera has been observed in the last decades. This pollinator plays a fundamental role in food production and the economy in Europe. The decline of honey bee colonies is linked to several stressors, including pesticides. The current pesticide risk assessment of honey bees in Europe focuses on lethal effects and lacks reflection on sublethal effects. A better understanding of the consequences that exposure to these chemicals has on honey bees is still needed. In this context, the aim of this European Food Risk Assessment Fellowship Programme fellowship project has been to use in silico methodologies, such as virtual screening, as a first step to identify possible interactions at the molecular level between A. mellifera proteins and pesticide ligands. For this purpose, a docking study of the proteins from A. mellifera and pesticide ligands extracted from online databases has been performed by using the software Autodock Vina. The results obtained were a ranking based on the predicted affinity of the pesticides for specific and non‐specific binding sites on bee macromolecules. These results were compared with data obtained from the literature and linked to potential sublethal effects. Finally, a risk assessment analysis of the identified molecular stressors of honey bees was performed. The results of this study are considered a starting point to identify new sources of possible stress for honey bees and thereby contribute to the overall understanding of the honey bee decline

Microbial Electrochemical Systems: Principles, Construction and Biosensing Applications

Microbial electrochemical systems are a fast emerging technology that use microorganisms to harvest the chemical energy from bioorganic materials to produce electrical power. Due to their flexibility and the wide variety of materials that can be used as a source, these devices show promise for applications in many fields including energy, environment and sensing. Microbial electrochemical systems rely on the integration of microbial cells, bioelectrochemistry, material science and electrochemical technologies to achieve effective conversion of the chemical energy stored in organic materials into electrical power. Therefore, the interaction between microorganisms and electrodes and their operation at physiological important potentials are critical for their development. This article provides an overview of the principles and applications of microbial electrochemical systems, their development status and potential for implementation in the biosensing field. It also provides a discussion of the recent developments in the selection of electrode materials to improve electron transfer using nanomaterials along with challenges for achieving practical implementation, and examples of applications in the biosensing field

Microbial Electrochemical Systems: Principles, Construction and Biosensing Applications

Microbial electrochemical systems are a fast emerging technology that use microorganisms to harvest the chemical energy from bioorganic materials to produce electrical power. Due to their flexibility and the wide variety of materials that can be used as a source, these devices show promise for applications in many fields including energy, environment and sensing. Microbial electrochemical systems rely on the integration of microbial cells, bioelectrochemistry, material science and electrochemical technologies to achieve effective conversion of the chemical energy stored in organic materials into electrical power. Therefore, the interaction between microorganisms and electrodes and their operation at physiological important potentials are critical for their development. This article provides an overview of the principles and applications of microbial electrochemical systems, their development status and potential for implementation in the biosensing field. It also provides a discussion of the recent developments in the selection of electrode materials to improve electron transfer using nanomaterials along with challenges for achieving practical implementation, and examples of applications in the biosensing field