Biochemical characterization and DNA binding properties of the MocR family member GabR, a PLP-dependent transcription factor

GabR è un fattore di trascrizione chimerico appartenente alla famiglia dei MocR/GabR, costituito da un dominio N-terminale elica-giro-elica di legame al DNA e un dominio effettore e/o di oligomerizzazione al C-terminale. I due domini sono connessi da un linker flessibile di 29 aminoacidi. Il dominio C-terminale è strutturalmente omologo agli enzimi aminotransferasici fold-type I, i quali, utilizzando il piridossal-5’-fosfato (PLP) come cofattore, sono direttamente coinvolti nel metabolismo degli aminoacidi. L’interazione contemporanea di PLP e acido γ-aminobutirrico (GABA) a GabR fa sì che questa promuova la trascrizione di due geni, gabT e gabD, implicati nel metabolismo del GABA. GabR cristallizza come un omodimero con una configurazione testa-coda. Il legame con la regione promotrice gabTD avviene attraverso il riconoscimento specifico di due sequenze dirette e ripetute (ATACCA), separate da uno spacer di 34 bp. In questo studio sono state indagate le proprietà biochimiche, strutturali e di legame al DNA della proteina GabR di Bacillus subtilis. L’analisi spettroscopica dimostra che GabR interagisce con il PLP formando l’aldimina interna, mentre in presenza di GABA si ottiene l’aldimina esterna. L’interazione fra il promotore gabTD e le forme holo e apo di GabR è stata monitorata mediante Microscopia a Forza atomica (AFM). In queste due condizioni di legame è stata stimata una Kd di circa 40 ηM. La presenza di GABA invece, determinava un incremento di circa due volte della Kd, variazioni strutturali nei complessi GabR-DNA e una riduzione del compattamento del DNA alla proteina, indipendentemente dalla sequenza del promotore in esame. Al fine di valutare il ruolo delle caratteristiche topologiche del promotore, sono state inserite cinque e dieci bp all’interno della regione spacer che separa le due sequenze ripetute dirette riconosciute da GabR. I significativi cambiamenti topologici riscontrati nel frammento aggiunto di cinque bp si riflettono anche sulla forte riduzione dell’affinità di legame verso la proteina. Al contrario, l’inserzione di 10 bp provoca solamente l’allontanamento delle sequenze ripetute dirette. L’assenza quindi di cambiamenti significativi nella topologia di questo promotore fa sì che l’affinità di legame per GabR rimanga pressoché inalterata rispetto al promotore non mutato. L’analisi del potenziale elettrostatico superficiale di GabR mostra la presenza di una fascia carica positivamente che si estende lungo un’intera faccia della proteina. Per verificare l’importanza di questa caratteristica di GabR nel meccanismo di interazione al DNA, sono stati preparati ed indagati i mutanti R129Q e K362-366Q, in cui la carica positiva superficiale risultava indebolita. L’affinità di legame dei mutanti di GabR per il DNA era inferiore rispetto alla proteina non mutata, in particolar modo nel mutante K362-366Q. Le evidenze acquisite suggeriscono che la curvatura intrinseca del promotore ed il corretto orientamento delle sequenze sulla doppia elica, più della distanza che le separa, siano critici per sostenere l’interazione con GabR. Oltre a questo, la superficie positiva di GabR è richiesta per accomodare la curvatura del DNA sul corpo della proteina. Alla luce di questo, l’interazione GabR-gabTD è un esempio di come il riconoscimento specifico di sequenze, la topologia del DNA e le caratteristiche strutturali della proteina siano contemporaneamente necessarie per sostenere un’interazione proteina-DNA stabile.

Spin dynamics of current-driven single magnetic adatoms and molecules

A scanning tunneling microscope can probe the inelastic spin excitations of a single magnetic atom in a surface via spin-flip assisted tunneling in which transport electrons exchange spin and energy with the atomic spin. If the inelastic transport time, defined as the average time elapsed between two inelastic spin flip events, is shorter than the atom spin-relaxation time, the scanning tunnel microscope (STM) current can drive the spin out of equilibrium. Here we model this process using rate equations and a model Hamiltonian that describes successfully spin-flip-assisted tunneling experiments, including a single Mn atom, a Mn dimer, and Fe Phthalocyanine molecules. When the STM current is not spin polarized, the nonequilibrium spin dynamics of the magnetic atom results in nonmonotonic dI/dV curves. In the case of spin-polarized STM current, the spin orientation of the magnetic atom can be controlled parallel or antiparallel to the magnetic moment of the tip. Thus, spin-polarized STM tips can be used both to probe and to control the magnetic moment of a single atom.

Storage of classical information in quantum spins

Digital magnetic recording is based on the storage of a bit of information in the orientation of a magnetic system with two stable ground states. Here we address two fundamental problems that arise when this is done on a quantized spin: quantum spin tunneling and backaction of the readout process. We show that fundamental differences exist between integer and semi-integer spins when it comes to both reading and recording classical information in a quantized spin. Our findings imply fundamental limits to the miniaturization of magnetic bits and are relevant to recent experiments where a spin-polarized scanning tunneling microscope reads and records a classical bit in the spin orientation of a single magnetic atom.

Characterization of rock slopes through slope mass rating using 3D point clouds

Rock mass classification systems are widely used tools for assessing the stability of rock slopes. Their calculation requires the prior quantification of several parameters during conventional fieldwork campaigns, such as the orientation of the discontinuity sets, the main properties of the existing discontinuities and the geo-mechanical characterization of the intact rock mass, which can be time-consuming and an often risky task. Conversely, the use of relatively new remote sensing data for modelling the rock mass surface by means of 3D point clouds is changing the current investigation strategies in different rock slope engineering applications. In this paper, the main practical issues affecting the application of Slope Mass Rating (SMR) for the characterization of rock slopes from 3D point clouds are reviewed, using three case studies from an end-user point of view. To this end, the SMR adjustment factors, which were calculated from different sources of information and processes, using the different softwares, are compared with those calculated using conventional fieldwork data. In the presented analysis, special attention is paid to the differences between the SMR indexes derived from the 3D point cloud and conventional field work approaches, the main factors that determine the quality of the data and some recognized practical issues. Finally, the reliability of Slope Mass Rating for the characterization of rocky slopes is highlighted.

SCAPHOID AND LUNATE CARPAL MECHANICS OVER THE SPECTRUM OF HEALTHY FUNCTION

When ligaments within the wrist are damaged, the resulting loss in range of motion and grip strength can lead to reduced earning potential and restricted ability to perform important activities of daily living. Left untreated, ligament injuries ultimately lead to arthritis and chronic pain. Surgical repair can mitigate these issues but current procedures are often non-anatomic and unable to completely restore the wrist’s complex network of ligaments. An inability to quantitatively assess wrist function clinically, both before and after surgery, limits the ability to assess the response to clinical intervention. Previous work has shown that bones within the wrist move in a similar pattern across people, but these patterns remain challenging to predict and model. In an effort to quantify and further develop the understanding of normal carpal mechanics, we performed two studies using 3D in vivo carpal bone motion analysis techniques. For the first study, we measured wrist laxity and performed CT scans of the wrist to evaluate 3D carpal bone positions. We found that through mid-range radial-ulnar deviation range of motion the scaphoid and lunate primarily flexed and extended; however, there was a significant relationship between wrist laxity and row-column behaviour. We also found that there was a significant relationship between scaphoid flexion and active radial deviation range of motion. For the second study, an analysis was performed on a publicly available database. We evaluated scapholunate relative motion over a full range of wrist positions, and found that there was a significant amount of variation in the location and orientation of the rotation axis between the two bones. Together the findings from the two studies illustrate the complexity and subject specificity of normal carpal mechanics, and should provide insights that can guide the development of anatomical wrist ligament repair surgeries that restore normal function.

Diritti sociali e crisi del costituzionalismo europeo [Social rights and the crisis of European constitutionalism]. WP C.S.D.L.E. “Massimo D’Antona”.INT – 95/2012

In the face of what looks like a real impasse of the the line of European constitutionalism, partly due to an orientation of the Court of Justice which tends to favor the protection of economic freedoms over the protection of social rights, two opposing trends occurr. The first amounts to a new "constitutional patriotism"; the second entrusts the protection of fundamental social rights no longer to a single Chart or to a single court but to a multi-level system of protection. A dialogue between the European courts that truly valorizes fundamental rights, however, might be hindered by what someone has seen as a resurgence of the dualist theories, evident in an ECJ’s decision as Kadi.

The Pro-Western Turn in Russia's Foreign Policy: Causes, Consequences and Prospects. OSW Point of View, September 2002

From Europe and Poland's point of view, one of the most important recent developments in international politics was the re-orientation of Russia's foreign policy. This paper aims to answer three important questions relating to this issue: 4. When and why did the "pro-Western turn" in the Russian Federation's policy take place? 5. Has it been profitable for Russia? 6. What goals will the Russian policy pursue in the future? An analysis of the last two years in Russia's foreign policy leads to the several conclusions, including those: 5. Clearly, the Russian leaders realise that in the longer term, Russia - in its desire for more influence in the world - will not be able to survive as an independent pole of power in international politics and it will have to join forces with the West (most likely, the European Union). 6. September 11 was not the cause of Russia's pro-Western turn, but rather a catalyst that put the process which started when Vladimir Putin took his office in sharp focus. 7. In the nearest future, this new direction of Russia's foreign policy seems not be challenged by internal opposition in Russia. 8. The "pro-Western turn" proved to be beneficial for Russia, although: d. Russia has not become a strategic ally of the US e. There has been no breakthrough in the relations between Russia and the European Union, and Moscow has not gained any real influence on NATO's important decisions. f. Russia has not become a major decision-maker of international politics. 5. Russia's closing to the West is in Poland's and Europe's interest.

The "Pro-Western Turn" in Russia's Foreign Policy: Causes, Consequences and Prospects. OSW Point of View (unspecified), September 2002

From Europe and Poland's point of view, one of the most important recent developments in international politics was the re-orientation of Russia's foreign policy. This paper aims to answer three important questions relating to this issue: 1. When and why did the "pro-Western turn" in the Russian Federation's policy take place? 2. Has it been profitable for Russia? 3. What goals will the Russian policy pursue in the future? An analysis of the last two years in Russia's foreign policy leads to the several conclusions, including those: a. Clearly, the Russian leaders realise that in the longer term, Russia - in its desire for more influence in the world - will not be able to survive as an independent pole of power in international politics and it will have to join forces with the West (most likely, the European Union). b. September 11 was not the cause of Russia's pro-Western turn, but rather a catalyst that put the process which started when Vladimir Putin took his office in sharp focus. 7. In the nearest future, this new direction of Russia's foreign policy seems not be challenged by internal opposition in Russia. c. The "pro-Western turn" proved to be beneficial for Russia, although: d. Russia has not become a strategic ally of the US e. There has been no breakthrough in the relations between Russia and the European Union, and Moscow has not gained any real influence on NATO's important decisions. f. Russia has not become a major decision-maker of international politics. g. Russia's closing to the West is in Poland's and Europe's interest.

The Effectiveness of Firms' Strategic Orientations: Investigating Multiple Theoretical Perspectives

The strategic orientation of firms can take on many forms. Researchers most commonly distinguish between entrepreneurial, market, and learning orientations. In combination, strategic orientations represent a firm's value proposition in terms of the markets in which it operates, where it deploys its resources, and which behavioral patterns are established. This thesis provides insights into the effectiveness of strategic orientations by adopting multiple theoretical perspectives. The strategic orientations of entrepreneurial, market, learning, and innovation orientations are investigated in an isolated as well as interrelated manner. The first research article concentrates on entrepreneurial orientation as its conceptualization and operationalization is subject to several debates in the literature. This conceptual study shows how the challenges of the entrepreneurial orientation construct can be overcome in future research to arrive at a higher level of construct clarity. Thereby, the theoretical perspectives of entrepreneurial dominant logic and the theory of planned behavior are employed. The literature has predominantly focused on investigating the effectiveness of particular strategic orientations. Recently, scholars have stressed their synergetic impact on firm performance and, as such, the relevance of considering their combined role in creating superior value for firms. However, empirical research on their interrelatedness remains scant and dispersed, making it necessary to conduct further research on strategic orientations in an integrative manner. As such, the second research article demonstrates which interrelated roles are played by entrepreneurial, market, and learning orientations in their relationship to firm performance. The rich body of existing knowledge is synthesized by means of meta-analysis under the perspective of strategic coalignment as well as the resource-based view of the firm.

The influence of reading direction on hemianopic reading disorders.

Hemianopic reading impairment is a consequence of a visual field defect to either the right or the left side and is characterized by an increased reading time and reduced reading performance. Depending on the side of the visual field defect, reading will be affected differently: Patients suffering from a visual field defect to the right side have noticeable difficulties in reading fluently with slowing. Patients suffering from a visual field defect to the left usually struggle to find the beginning of a line and read more fluently. It was suggested in the literature that changing the reading direction from horizontal to vertical may be a training strategy to reduce reading problems in patients with hemianopia. The aim of the study was to investigate the influence of reading direction on reading speed in patients with left- or right-sided visual field defects and in healthy controls. METHOD In 13 patients with hemianopia and in 13 age-matched controls, reading speed was calculated for texts in standard as well as in clockwise rotated orientation of 90, 180, and 270°. RESULTS In both groups, text rotation reduced reading speed compared to standard reading. Patients with left-sided hemianopia had the greatest reduction after text rotation. Patients with right-sided hemianopia had the smallest speed reduction in 90° vertically rotated texts. CONCLUSIONS Text rotation has different effects in left- or right-sided hemianopia patients. For patients with left-sided heminanopia, rotation of the text may not be a helpful training strategy, for right-sided hemianopia vertical rotation of the text of 90° may be a beneficial training strategy to reduce reading deficits.

Sculpture of Portia over front door of Martha Cook Residence Hall

Source image was printed BACKWARDS. 2 scans made, 1of2 "as is", 2of2 reversed in photoshop for correct orientation of sculpture

RESEARCHING SITUATED LEARNING AS THE DEVELOPMENT OF PRACTICE FOR COLLEGE ACCESS PRACTITIONERS: A CASE STUDY OF THE COLLEGE ASSISTANCE MIGRANT PROGRAM

Thesis (Ph.D.)--University of Washington, 2016-03

Microanalysis of epitaxially grown diamond tip array

Electron backscattering diffraction has been applied on polycrystalline diamond films grown using microwave plasma assisted chemical vapour deposition on silicon substrate, in order to provide a map of the individual diamond grains, grain boundary, and the crystal orientation of discrete crystallites. The nucleation rate and orientation are strongly affected by using a voltage bias on the substrate to influence and enhance the nucleation process, the bias enhanced nucleation process. In this work, the diamond surface is mapped using electron backscattering diffraction, then a layer of a few microns is ion milled away exposing a lower layer for analysis and so on. This then permits a three dimensions reconstruction of the film texture.

The 3-D Structure of Polycrystalline Diamond Film by Electron Backscattering Diffraction (EBSD)

A lithographic method was used to produce polycrystalline diamond films having highly defined surface geometry, showing an array of diamond tips for possible application as a field emitter device. The films grown in this study used microwave plasma assisted chemical vapour deposition (MACVD) on a silicon substrate; the substrate was then dissolved away to reveal the surface features on the diamond film. It is possible to align the crystallite direction and affect the electron emission properties using a voltage bias to enhance the nucleation process and influence the nuclei to a preferred orientation. This study focuses on the identification of the distribution of crystal directions in the film, using electron backscattering diffraction (EBSD) to identify the crystallographic character of the film surface. EBSD allows direct examination of the individual diamond grains, grains boundaries and the crystal orientation of each individual crystallite. The EBSD maps of the bottom (nucleation side) of the films, following which a layer of film is ion-milled away and the mapping process repeated. The method demonstrates experimentally that oriented nucleation occurs and the thin sections allow the crystal texture to be reconstructed in 3-D. (C) 2003 Elsevier B.V. All rights reserved.

Voltage control of exchange coupling in phosphorus doped silicon

Motivated by applications to quantum computer architectures we study the change in the exchange interaction between neighbouring phosphorus donor electrons in silicon due to the application of voltage biases to surface control electrodes. These voltage biases create electro-static fields within the crystal substrate, perturbing the states of the donor electrons and thus altering the strength of the exchange interaction between them. We find that control gates of this kind can be used to either enhance or reduce the strength of the interaction, by an amount that depends both on the magnitude and orientation of the donor separation.