Cytochrome P450(cin) (CYP176A), isolation, expression, and characterization

Cytochromes P450 are members of a superfamily of hemoproteins involved in the oxidative metabolism of various physiologic and xenobiotic compounds in eukaryotes and prokaryotes. Studies on bacterial P450s, particularly those involved in monoterpene oxidation, have provided an integral contribution to our understanding of these proteins, away from the problems encountered with eukaryotic forms. We report here a novel cytochrome P450 (P450(cin), CYP176A1) purified from a strain of Citrobacter braakii that is capable of using cineole 1 as its sole source of carbon and energy. This enzyme has been purified to homogeneity and the amino acid sequences of three tryptic peptides determined. By using this information, a PCR-based cloning strategy was developed that allowed the isolation of a 4-kb DNA fragment containing the cytochrome P450(cin) gene (cinA). Sequencing revealed three open reading frames that were identified on the basis of sequence homology as a cytochrome P450, an NADPH-dependent flavodoxin/ferrodoxin reductase, and a flavodoxin. This arrangement suggests that P450(cin) may be the first isolated P450 to use a flavodoxin as its natural redox partner. Sequencing also identified the unprecedented substitution of a highly conserved, catalytically, important active site threonine with an asparagine residue. The P450 gene was subcloned and heterologously expressed in Escherichia coli at similar to2000 nmol/liter of original culture, and purification was achieved by standard protocols. Postulating the native E. coli flavodoxin/flavodoxin reductase system might mimic the natural redox partners of P450,in, it was expressed in E. coli in the presence of cineole 1. A product was formed in vivo that was tentatively identified by gas chromatography-mass spectrometry as 2-hydroxycineole 2. Examination of P450(cin) by UV-visible spectroscopy revealed typical spectra characteristic of P450s, a high affinity for cineole 1 (K-D = 0.7 mum), and a large spin state change of the heme iron associated with binding of cineole 1. These facts support the hypothesis that cineole 1 is the natural substrate for this enzyme and that P450(cin) catalyzes the initial monooxygenation of cineole 1 biodegradation. This constitutes the first characterization of an enzyme involved in this pathway.

Financiamento dos Sistema de Saude

Este Relatório sobre a Saúde no Mundo foi produzido sob a direcção geral de Carissa Etienne, Assistente do Director-Geral, Sistemas e Serviços de Saúde e Anarfi Asamoa-Baah, Director Geral Adjunto. Os redactores principais froam David B Evans, Riku Elovainio e Gary Humphreys; com contribuições de Daniel Chisholm, Joseph Kutzin, Sarah Russell, Priyanka Saksena e Ke Xu. Contribuições sob a forma de caixas de texto e análises foram fornecidos por: Ole Doetinchem, Adelio Fernandes Antunes, Justine Hsu, Chandika K. Indikadahena, Jeremy Lauer, Nathalie van de Maele, Belgacem Sabri, Hossein Salehi, Xenia Scheil-Adlung (ILO) and Karin Stenberg. Sugestões e comentários foram recebidos dos Directores Regionais, Assistentes do Director-Geral e respectivas equipas. Análises, dados e revisões da organização do texto, vários rascunhos ou secções específicas foram fornecidos por (em adição às pessoas jáacima mencionadas): Dele Abegunde, Michael Adelhardt, Hector Arreola, Guitelle Baghdadi-Sabeti, Dina Balabanova, Dorjsuren Bayarsaikhan, Peter Berman, Melanie Bertram, Michael Borowitz, Reinhard Busse, Alexandra Cameron, Guy Carrin, Andrew Cassels, Eleonora Cavagnero, John Connell, David de Ferranti, Don de Savigny, Varatharajan Durairaj, Tamás Evetovits, Josep Figueras, Emma Fitzpatrick, Julio Frenk, Daniela Fuhr, Ramiro Guerrero, Patricia Hernandez Pena, Hans V Hogerzeil, Kathleen Holloway, Melitta Jakab, Elke Jakubowski, Christopher James, Mira Johri, Matthew Jowett, Joses Kirigia, Felicia Knaul, Richard Laing, Nora Markova, Awad Mataria, Inke Mathauer, Don Matheson, Anne Mills, Eduardo Missoni, Laurent Musango, Helena Nygren-Krug, Ariel Pablos-Mendez, Anne-Marie Perucic, Claudia Pescetto, Jean Perrot, Alexander Preker, Magdalena Rathe, Dag Rekve, Ritu Sadana, Rocio Saenz, Thomas Shakespeare, Ian Smith, Peter C Smith, Alaka Singh, Ruben Suarez Berenguela, Tessa Tan-Torres Edejer, Richard Scheffler, Viroj Tangcharoensathien, Fabrizio Tediosi, Sarah Thomson, Ewout van Ginneken, Cornelis van Mosseveld e Julia Watson. A redacção do Relatório foi informada por muitos indivíduos de várias instituições que forneceram documentos de suporte; estes documentos de suporte podem ser encontrados em: http://www.who.int/healthsystems/topics/financing/healthreport/whr_background/en Michael Reid editou as cópias do Relatório, Gaël Kernen produziu as figuras e Evelyn Omukubi forneceu o valioso apoio secretarial e administrativo. O desenho e paginação foi feito por Sophie Guetaneh Aguettant e Cristina Ortiz. Ilustração por Edel Tripp (http://edeltripp.daportfolio.com). A tradução foi realizada por Jorge Cabral e Aurélio Floriano e revista por Aurélio Floriano e Paulo Ferrinho, do Instituto de Higiene e Medicina Tropical, da Universidade Nova de Lisboa - Lisboa, Portugal. A publicação foi produzida com o apoio da Comunidade dos Países de Língua Portuguesa (CPLP), sob autorização do Director Geral da Organização Mundial da Saúde (OMS). As informações contidas neste Relatório não podem, de forma alguma, ser tomadas como a expressão das posições da CPLP

Counteraction of HLA-C-mediated immune control of HIV-1 by Nef.

A host genetic variant (-35C/T) correlates with increased human leukocyte antigen C (HLA-C) expression and improved control of HIV-1. HLA-C-mediated immunity may be particularly protective because HIV-1 is unable to remove HLA-C from the cell surface, whereas it can avoid HLA-A- and HLA-B-mediated immunity by Nef-mediated down-modulation. However, some individuals with the protective -35CC genotype exhibit high viral loads. Here, we investigated whether the ability of HIV-1 to replicate efficiently in the "protective" high-HLA-C-expression host environment correlates with specific functional properties of Nef. We found that high set point viral loads (sVLs) were not associated with the emergence of Nef variants that had acquired the ability to down-modulate HLA-C or were more effective in removing HLA-A and HLA-B from the cell surface. However, in individuals with the protective -35CC genotype we found a significant association between sVLs and the efficiency of Nef-mediated enhancement of virion infectivity and modulation of CD4, CD28, and the major histocompatibility complex class II (MHC-II)-associated invariant chain (Ii), while this was not observed in subjects with the -35TT genotype. Since the latter Nef functions all influence the stimulation of CD4(+) T helper cells by antigen-presenting cells, they may cooperate to affect both the activation status of infected T cells and the generation of an antiviral cytotoxic T-lymphocyte (CTL) response. In comparison, different levels of viremia in individuals with the common -35TT genotype were not associated with differences in Nef function but with differences in HLA-C mRNA expression levels. Thus, while high HLA-C expression may generally facilitate control of HIV-1, Nef may counteract HLA-C-mediated immune control in some individuals indirectly, by manipulating T-cell function and MHC-II antigen presentation.

Magnetism of isolated Mn12 single-molecule magnets detected by magnetic circular dichroism: Observation of spin tunneling with a magneto-optical technique

We report magnetic and magneto-optical measurements of two Mn12 single-molecule magnet derivatives isolated in organic glasses. Field-dependent magnetic circular dichroism (MCD) intensity curves (hysteresis cycles) are found to be essentially identical to superconducting quantum interference device magnetization results and provide experimental evidence for the potential of the optical technique for magnetic characterization. Optical observation of magnetic tunneling has been achieved by studying the decay of the MCD signal at weak applied magnetic field

Why not all young firms invest in R&D

This article aims to analyze the different impact that some factors may exert on the probability that a small young firm invests intensively in R&D. Recently, an increasing amount of the literature makes reference to the vital role played by a small number of young firms in generating jobs and increasing efficiency levels. However, not all new firms invest in R&D. Departing from the definition of YICs (firms younger than 6 years old, fewer than 250 employees and with more than 15% of their revenues invested in R&D activities), and with an extensive sample of the Spanish Community Innovation Survey between 2004- 2010, we try to determine: i) those factors that cause firms to become YICs (innovative young small firms) or YNICs (moderately innovative young small firms); ii) what is the difference in the impact of those factors between YICs and YNICs. Our results show that factors such as initial innovation capacity and cooperation in R&D projects enhance the probability of becoming a YIC. Nevertheless, factors such as export potential and market uncertainty may influence the decision to invest moderately and become a YNIC. Keywords: Innovation, Policy, YICs. JEL Classifications: O31, D21

Assessment of inflammatory Bowel Disease patient's needs and problems from a nursing perspective

BACKGROUND: In this study, we aimed at assessing Inflammatory Bowel Disease patients' needs and current nursing practice to investigate to what extent consensus statements (European Crohn's and Colitis Organization) on the nursing roles in caring for patients with IBD concur with local practice. METHODS: We used a mixed-method convergent design to combine quantitative data prospectively collected in the Swiss IBD cohort study and qualitative data from structured interviews with IBD healthcare experts. Symptoms, quality of life, and anxiety and depression scores were retrieved from physician charts and patient self-reported questionnaires. Descriptive analyses were performed based on quantitative and qualitative data. RESULTS: 230 patients of a single center were included, 60% of patients were males, and median age was 40 (range 18-85). The prevalence of abdominal pain was 42%. Self-reported data were obtained from 75 out of 230 patients. General health was perceived significantly lower compared with the general population (p < 0.001). Prevalence of tiredness was 73%; sleep problems, 78%; issues related to work, 20%; sexual constraints, 35%; diarrhea, 67%; being afraid of not finding a bathroom, 42%; depression, 11%; and anxiety symptoms, 23%. According to experts' interviews, the consensus statements are found mostly relevant with many recommendations that are not yet realized in clinical practice. CONCLUSION: Identified prevalence may help clinicians in detecting patients at risk and improve patient management. © 2015 S. Karger AG, Basel.

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Sosiaalisen liikkuvuuden tutkimusta yhdistämässä - tai sitten ei

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Frailty, an Emerging Research and Clinical Paradigm: Issues and Controversies

Ne pas citer. Version pour diffusion uniquement. Citer l'article une fois publié. / Not to be cited. For distribution only. Cite article once published.

Magnetism of isolated Mn12 single-molecule magnets detected by magnetic circular dichroism: Observation of spin tunneling with a magneto-optical technique

We report magnetic and magneto-optical measurements of two Mn12 single-molecule magnet derivatives isolated in organic glasses. Field-dependent magnetic circular dichroism (MCD) intensity curves (hysteresis cycles) are found to be essentially identical to superconducting quantum interference device magnetization results and provide experimental evidence for the potential of the optical technique for magnetic characterization. Optical observation of magnetic tunneling has been achieved by studying the decay of the MCD signal at weak applied magnetic field