1,720,957 research outputs found
IMP-GMP specific cytosolic 5′-nucleotidase regulates nucleotide pool and prodrug metabolism
No full textBackground Type II cytosolic 5′-nucleotidase (cN-II) catalyzes the hydrolysis of purine and, to some extent, of pyrimidine monophosphates. Recently, a number of papers demonstrated the involvement of cN-II in the mechanisms of resistance to antitumor drugs such as cytarabine, gemcitabine and fludarabine. Furthermore, cN-II is involved in drug resistance in patients affected by hematological malignancies influencing the clinical outcome. Although the implication of cN-II expression and/or activity appears to be correlated with drug resistance and poor prognosis, the molecular mechanism by which cN-II mediates drug resistance is still unknown. Methods HEK 293 cells carrying an expression vector coding for cN-II linked to green fluorescent protein (GFP) and a control vector without cN-II were utilized. A highly sensitive capillary electrophoresis method was applied for nucleotide pool determination and cytotoxicity exerted by drugs was determined with 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay. Results Over-expression of cN-II causes a drop of nucleoside triphosphate concentration and a general disturbance of nucleotide pool. Over-expressing cells were resistant to fludarabine, gemcitabine and cytarabine independently of cN-II ability to hydrolyze their monophosphates. Conclusions An increase of cN-II expression is sufficient to cause both a general disturbance of nucleotide pool and an increase of half maximal inhibitory concentration (IC50) of the drugs. Since the monophosphates of cytarabine and gemcitabine are not substrates of cN-II, the protection observed cannot be directly ascribed to drug inactivation. General significance Our results indicate that cN-II exerts a relevant role in nucleotide and drug metabolism through not only enzyme activity but also a mechanism involving a protein-protein interaction, thus playing a general regulatory role in cell survival. Sentence Resistance to fludarabine, gemcitabine and cytarabine can be determined by an increase of cN-II both through dephosphorylation of active drugs and perturbation of nucleotide pool
A native electrophoretic technique to study oligomerization and activity of cytosolic 5'-nucleotidase II
No full textStructural and functional studies of cytosolic 5’-nucleotidase II (cN-II)
No full textABSTRACT
5’-Nucleotidases, which dephosphorylate non-cyclic (deoxy)ribonucleoside monophosphates to nucleosides and orthophosphate, constitute a heterogeneous family of widespread enzymes that vary in subcellular location, tissue-specific expression and substrate specificity.
Seven human 5’-nucleotidases have been isolated, five of which are located in the cytosol (cN-IA, cN-IB, cN-II, cN-II, cdN), one in the mitochondrial matrix (mdN) and one is anchorated to the outer plasma membrane by a glycosyl phosphatidylinositol anchor (eN). By opposing the phosphorylation of nucleosides by kinases, intracellular 5’-nucleotidases take part in substrate cycles thereby regulating the intracellular levels of (deoxy)ribonucleoside pools.
The focus of this research project is the cytosolic 5’-nucleotidase II (cN-II), a widespread enzyme with a remarkable sequence conservation through evolution. The Mg2+-dependent cN-II is a bifunctional enzyme, since it can transfer the orthophosphate from a nucleoside monophosphate (preferentially (d)IMP and (d)GMP) not only to water, but also to the 5’ position of a nucleoside acceptor, usually (d)inosine, by a ping-pong reaction mechanism; this is due to the formation of a pentacovalent phosphoester enzyme intermediate, a feature of all members of the HAD superfamily cN-II belongs to. CN-II is submitted to an intricate allosteric regulation, being activated by several phosphorylated compounds (Ap4A, dATP, ATP, ADP, 2,3-BPG) and inhibited by inorganic phosphate; moreover adenylate energy charge is able to affect its activity. In particular, at physiological values of adenylate energy charge, pH and Pi, if a suitable nucleoside acceptor is available, cN-II behaves mainly as a phosphotransferase. Nevertheless, it is unknown whether the transfer of orthophosphate normally occurs in vivo, as Km values for nucleoside substrates are quite high. In vivo, in fact, hydrolysis seems to be cN-II major activity.
CN-II has always been described as a homotetramer of 60 kDa subunits: the crystallized structure of cN-II highlighted the residues responsible for the interaction of two single subunits at interface A and those which hold the two dimers together at interface B. Besides, also the presence of two putative effector sites has been hypothesized. Moreover, it has been proposed by other authors that the effectors influence cN-II oligomerization state: the activators would promote subunit aggregation, while Pi woud act in the opposite way.
Clinical interest in this enzyme increases enormously in these last years, as it seems that cN-II is involved in anticancer and antiviral prodrug metabolism: in fact, not only it could be responsible for resistance to nucleoside analogues with its catabolic function, but it could also be able to activate them by its phosphotransferase activity. Besides mRNA level of cN-II probably has a prognostic value in adult acute myeloid leukemia.
In the light of this, any findings concerning how cN-II activity is regulated and its physiological role will offer new possible therapeutical approaches. Therefore, we designed this research project combining two experimental branches: one focused on the regulatory features of cN-II, seeking to ascertain if the effectors are able to affect cN-II subunit association and enzymatic activity; the other, instead, directed towards the clarification of the role of cN-II in purine metabolism.
In this way, the results obtained would help to improve existing therapeutic treatments leading to the design of personalized chemotherapy; moreover, the cellular systems we built could be very important for the determination not only of the role of cN-II in purine metabolism but also of the relationship between its altered activity and neurological and/or neoplastic pathologies
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Variations on the Author
Full text link“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
Appropriate Similarity Measures for Author Cocitation Analysis
Full text linkWe provide a number of new insights into the methodological discussion about author cocitation analysis. We first argue that the use of the Pearson correlation for measuring the similarity between authors’ cocitation profiles is not very satisfactory. We then discuss what kind of similarity measures may be used as an alternative to the Pearson correlation. We consider three similarity measures in particular. One is the well-known cosine. The other two similarity measures have not been used before in the bibliometric literature. Finally, we show by means of an example that our findings have a high practical relevance.information science;Pearson correlation;cosine;similarity measure;author cocitation analysis
Dispelling the Myths Behind First-author Citation Counts
Full text linkWe conducted a full-scale evaluative citation analysis study of scholars in the XML research field to explore just how different from each other author rankings resulting from different citation counting methods actually are, and to demonstrate the capability of emerging data and tools on the Web in supporting more realistic citation counting methods. Our results contest some common arguments for the continued
use of first-author citation counts in the evaluation of scholars, such as high correlations between author rankings by first-author citation counts and other citation
counting methods, and high costs of using more realistic citation counting methods that are not well-supported by the ISI databases. It is argued that increasingly available digital full text research papers make it possible for citation analysis studies to go beyond what the ISI databases have directly supported and to employ more
sophisticated methods
koamabayili/VECTRON-author-checklist: VECTRON author checklist
No full textWe have done our best to complete the author checklist relating to the use of animals in the hut study. Note that the objective for the hut study was to evaluate the IRS treatment applications for residual efficacy against Anopheles mosquitoes, including the local An. coluzzii mosquito population. Cows were only used to attract mosquitoes into the huts and no tests were carried out directly on the cows. The author checklist is intended for use with studies where experiments are carried out on animals, which is why we have had such difficulty in completing this for the hut study, as many of the questions do not relate to how the cows were used
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