1,721,171 research outputs found
Going Beyond Counting First Authors in Author Co-citation Analysis
Get PDFThe 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
Functional Analysis of the Ninth Subunit of Yeast RNA Polymerase II, RPB9
No full textRNA polymerase II is the eukaryotic enzyme that synthesizes mRNA. It is a complex enzyme that is highly regulated by many protein factors throughout transcription. RNA polymerase II comprises twelve subunits, each of which likely plays a specific role in the function of the enzyme. The ninth subunit, RPB9, is involved in the initiation and elongation stages of transcription. In yeast, deletion of RPB9 results in sensitivity to high and low temperatures, as well as to the drug 6-aza-uracil. RPB9-deficient strains exhibit an alteration in the selection of transcript start sites, with an upstream shift of the start site window. The RNA polymerase II isolated from RPB9-null strains is unable to recognize intrinsic pause sites during elongation. The polymerase molecules that do arrest cannot resume transcribing upon stimulation by the elongation factor TFIIS. RPB9 is a small polypeptide, containing two zinc-binding regions (Zn 1 and Zn2) connected by an intervening sequence (linker region). This study presents the results of a mutational analysis designed to assisn th various functions of RPB9 to regions within the subunit. The Zn1 region was shown to be required for the start site selection activity of RPB9. When expressed in an RPB9-null yeast strain, the Zn1 region was found to be sufficient to restore start site selections to the wildtype pattern. Using a gel shift assay and purified proteins, an RNA polymerase II binding region was located to a conserved sequence (D---DPTLPR) within the linker region. The elongation activity of RPB9 was ascribed to the Zn2 region. Conserved charged residues within Zn2 were essential for the ability of RPB9 to mediate the reactivation of RNA polymerase II upon stimulation by TFIIS. This region of Zn2 is analogous to a charged flexible loop within the zinc ribbon domain of TFIIS. These observations provide the basis for a preliminary model of RPB9 interactions with RNA polymerase II.Doctor of Philosophy (PhD
Variations on the Author
Get PDF“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
Structural Analysis of Macrocyclic Peptides bound to Huntingtin
Get PDFHuntington's Disease (HD) is a neurodegenerative disorder characterized by motor dysfunction, cognitive decline, and psychiatric disturbances, driven by an expanded polyglutamine tract in the huntingtin (HTT) protein. Despite extensive research, understanding HTT has been limited by the scarcity of high-quality reagents. Using the RaPID method, our lab identified six macrocycles with high affinity for HTT and/or HTT-HAP40 complexes, confirmed by surface plasmon resonance spectroscopy. Four macrocycles (HHL1, HHD3, HD4, and HL2) were found to bind and affinity purify HTT and/or HTT-HAP40 from cell extracts. Hydrogen-deuterium exchange mass spectrometry (HDX-MS) revealed binding pockets, guiding CryoEM for structural elucidation. In my work, I modeled, refined and analyzed cryo-EM structures of the protein-ligand complexes. The macrocycles bound to deep, conserved pockets in the HTT-HAP40 complex. I hypothesize that identified macrocycles target functionally important regions and that they can be used as tools to unravel the function of these pockets.M.Sc
Appropriate Similarity Measures for Author Cocitation Analysis
Get PDFWe 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
Structural Basis for Enzyme Promiscuity and Specificty - Insights from Human Cytosolic sulfotransferase (SULT) and Sirtuin (SIRT) Families
Get PDFUnderstanding the structural basis of specificity and promiscuity of paralogous enzymes is important for deciphering molecular mechanisms and is a necessary step towards designing enzyme-specific modulators. The main objective of this thesis is to provide structural insights that relate protein local sequences to their observed binding and activity profiles through the study of two human protein families – cytosolic sulfotransferases (SULTs) and sirtuins (SIRTs). This was achieved by comparing the family-wide ligand binding fingerprints of these two enzyme families with the structural details of their corresponding enzyme-ligand co-crystal structures.
The hSULT enzyme family was profiled against a focused library through binding and activity assays. This suggested a number of novel compounds that bind to the less well-characterized SULT members (SULT1C3 and SULT4A1), and revealed additional broad-spectrum hSULT inhibitors. Based on the profiling data, three enzyme/co-factor/ligand complex structures were solved using X-ray crystallography. The structure of SULT1C2•PAP(3'-phosphoadenosine 5'-phosphate)•pentacholorphenol(PCP) provided a rationale for a novel SULTs inhibition mechanism that depends on substrate acidity. The SULT1B1•PAP•resveratrol structure suggested that the hydrogen-bonding coordination of the 5-OH group on resveratrol is the structural determinant for the observed substrate preference towards resveratrol. SULT2A1•PAP•lithocholic acid(LCA) ternary complex structure confirms that the specificity of SULT2A1 for lithocholic acid derives from its high hydrophobicity in the substrate binding pocket.
The same approach was used to interrogate the interaction of the sirtuins with their peptide substrates. The binding and enzymatic assays for human sirtuins have suggested that SIRT1 and SIRT2 are generally less discriminate against substrates while class IV sirtuins - SIRT6 and SIRT7 might be highly specific enzymes. Three different biochemical and kinetic assays showed that SIRT6-dependent histone deacetylation is about 1,000 times slower than for other highly active sirtuins. To understand the molecular basis for the specificity and low activity of SIRT6, I determined the first set of crystal structures for SIRT6 in complex with ADPr (ADP ribose) and the non-hydrolyzable analog of OAADPr (2’-O-acetyl-ADP ribose) – NAADPr (2’-N-acetyl-ADP ribose). The structures revealed human SIRT6 has unique structural features including a splayed zinc-binding domain, lacks a helix bundle and the conserved, highly flexible, NAD(+)-binding loop, which contribute to its observed biochemical behavior.Ph
Activity-based Functional Annotation of Unknown Proteins: HAD-like hydrolases from E. coli and S. cerevisiae
Get PDFIn all sequenced genomes, a large fraction of predicted genes encodes proteins of unknown biochemical function and up to 15% of the genes with ‘‘known’’ function are mis-annotated. Several global approaches are being employed to predict function, including sequence similarity searches, analysis of gene expression, protein interaction, and protein structure. Enzymes comprise a group of target proteins that require experimental characterization for accurate functional annotations. Here I applied enzyme genomics to identify new enzymes by screening individually purified proteins for enzymatic activity under relaxed reaction conditions, which allowed me to identify the subclass or sub-subclasses of enzymes to which the unknown protein belongs. Further biochemical characterization of proteins was facilitated by the application of secondary screens with natural substrates (substrate profiling). Application of general enzymatic screens and substrate profiling greatly sped up the identification of biochemical function of unknown proteins and the experimental verification of functional predictions produced by other functional genomics approaches.
As a test case, I used this approach to characterize the members of the haloacid dehalogenase (HAD)-like hydrolase superfamily, which consists mainly of uncharacterized enzymes, with a few members shown to possess phosphatase, beta-phosphoglucomutase, phosphonatase, and dehalogenase activities. Low sequence similarity between the members of the HAD superfamily precludes the computational prediction of their substrates and functions. Using a representative set of 80 phosphorylated substrates I characterized the phosphatase activities of 21 soluble HADs from Escherichia coli and seven soluble HADs from Saccharomyces cerevisiae. E. coli HADs show broad and overlapping substrate specificity against a wide range of phosphorylated metabolites. The yeast enzymes were more specific, and one protein also showed protein phosphatase activity. Comparison of HAD substrate profiles from two model organisms showed several “functional niches” that are occupied by HADs, which include hydrolysis of nucleotides, phosphoglycolate, phosphoserine, and pyridoxal phosphate. I proposed the cellular function for a number of HADs from both organisms based on substrate specificities. The physiological relevance of the phosphatase activity with the preferred substrate was validated in vivo for one of the HADs, E. coli YniC.Ph
Activity-based Functional Annotation of Unknown Proteins: HAD-like hydrolases from E. coli and S. cerevisiae
No full textIn all sequenced genomes, a large fraction of predicted genes encodes proteins of unknown biochemical function and up to 15% of the genes with ‘‘known’’ function are mis-annotated. Several global approaches are being employed to predict function, including sequence similarity searches, analysis of gene expression, protein interaction, and protein structure. Enzymes comprise a group of target proteins that require experimental characterization for accurate functional annotations. Here I applied enzyme genomics to identify new enzymes by screening individually purified proteins for enzymatic activity under relaxed reaction conditions, which allowed me to identify the subclass or sub-subclasses of enzymes to which the unknown protein belongs. Further biochemical characterization of proteins was facilitated by the application of secondary screens with natural substrates (substrate profiling). Application of general enzymatic screens and substrate profiling greatly sped up the identification of biochemical function of unknown proteins and the experimental verification of functional predictions produced by other functional genomics approaches.
As a test case, I used this approach to characterize the members of the haloacid dehalogenase (HAD)-like hydrolase superfamily, which consists mainly of uncharacterized enzymes, with a few members shown to possess phosphatase, beta-phosphoglucomutase, phosphonatase, and dehalogenase activities. Low sequence similarity between the members of the HAD superfamily precludes the computational prediction of their substrates and functions. Using a representative set of 80 phosphorylated substrates I characterized the phosphatase activities of 21 soluble HADs from Escherichia coli and seven soluble HADs from Saccharomyces cerevisiae. E. coli HADs show broad and overlapping substrate specificity against a wide range of phosphorylated metabolites. The yeast enzymes were more specific, and one protein also showed protein phosphatase activity. Comparison of HAD substrate profiles from two model organisms showed several “functional niches” that are occupied by HADs, which include hydrolysis of nucleotides, phosphoglycolate, phosphoserine, and pyridoxal phosphate. I proposed the cellular function for a number of HADs from both organisms based on substrate specificities. The physiological relevance of the phosphatase activity with the preferred substrate was validated in vivo for one of the HADs, E. coli YniC.Ph
Dispelling the Myths Behind First-author Citation Counts
Get PDFWe 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
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