Revisiting Allostery In Lac Repressor

Lac repressor (LacI) is an allosterically regulated transcription factor which controls expression of the lac operon in bacteria. LacI consists of a DNA-binding domain (DBD) and regulatory domain (RD), connected by a linker called the “hinge”. Binding of a small molecule inducer to the RD relieves repression through what is presumed to be a series of conformational changes mediated through the hinge. Despite decades of study, our understanding of this allosteric transition remains incomplete—mostly inferred from partial crystal structures and low-resolution scattering studies. In principle, solution-NMR could provide structural and dynamical information unobtainable by X-ray methods. However, due to LacI’s high molecular weight, low solubility, and transient stability, such studies have been limited to the non-allosteric, isolated DBD. Here, we present a solution-NMR study of the changes in structure and dynamics that underlie the allosteric transition of intact LacI. First, an optimized expression system is presented which enables characterization of LacI using NMR methodologies for high molecular weight proteins. Next, alternative NMR data sampling methods are implemented and further extended to overcome the low-solubility and transient stability limitations. Finally, these developments are combined to characterize LacI in each of its functional states. It is shown that the RD but not the DBD of apo LacI exists in an equilibrium between induced and repressed states with exchange occurring on the �s-ms timescale. Inducer binding in the absence of operator mostly quenches exchange but does not result in structural changes in the hinge or DBD. Conformational dynamics detected in the induced state are shown to be localized to a “network” of RD residues previously characterized to be critical for allostery. These dynamics are shown to be quenched in non-allosteric mutants which suggests functional relevance. Operator binding results in globally quenched dynamics and dramatic changes to the structure of the hinge. Inducer binding in the presence of operator results in only minor structural perturbation in the hinge and DBD. However, dynamics are shown to be activated in the RD. These results suggest that conformational dynamics may be critical to the allosteric transition of LacI

Inspired Design: Using Interdisciplinarity And Biomimicry For Software Innovation

This thesis presents research and proposes a framework for increasing the breadth and depth of interdisciplinary knowledge in the field of computer science. The intent is to address an increasing problem of complexity in software and computing systems. The approach is to equip software developers and computer scientists with a contextual perspective and a set of strategies for injecting innovation and creativity into the solutions they design by leveraging knowledge and models outside the traditional realm of computer science. A review of current and historical forms of interdisciplinarity and biomimicry are presented to build that context. The strategies presented include interdisciplinary education, interdisciplinary collaboration, interdisciplinary tools, biomimetic design, and the creation of new pattern languages based on nature\u27s design solutions. Each of these strategies stems from and leads to an open exchange of knowledge across disciplinary boundaries. When taken together, the knowledge and strategies presented are intended to inspire and foster a paradigm that recognizes the value of human and natural diversity as a source of innovation

Hemispheric contrasts of ice formation in stratiform supercooled liquid clouds: Long-term observations with the ground-based remote-sensing supersite LACROS

Die vorliegende Arbeit untersucht hemisphärische Unterschiede der heterogenen Eisbildung in unterkühlten Schichtwolken auf Basis von drei Datensätzen, die mit der mobilen bodengebundenen Fernerkundungsplattform LACROS (Leipzig Aerosol and Cloud Remote Observations System; Leipziger Aerosol- und Wolken- Fernerkundungssystem) erhoben wurden. Für die Nordhemisphäre wurden zwischen 2014 und 2018 gesammelte LACROS-Datensätze aus Leipzig (Deutschland, 51,4°N, 12,4°E) und Limassol (Zypern, 34,7°N, 33,0°E) verwendet. Ein zentraler Bestandteil dieser Arbeit war die Umsetzung des mehr als zwei Jahre umfassenden Einsatzes von LACROS im Rahmen der Kampagne DACAPO-PESO (Dynamics Aerosol Clouds And Precipitation Observation in the Pristine Environment of the Southern Ocean; Beobachtung von Dynamik, Aerosol, Wolken und Niederschlag in der unverschmutzen Umgebung des Südozeans) in Punta Arenas, Chile (53,1°S, 70,9°W). Dieser Datensatz stellt die ersten mehrjährigen bodengebundenen Fernerkundungsbeobachtungen in der westlichen Hälfte des Südozeans dar. Durch die Kombination aus Radar- und Lidarinstrumenten, einschließlich der Fähigkeit Vertikalbewegungen zu beobachten, ist es möglich, mit LACROS Aerosol-Wolken-Dynamik-Wechselwirkungen detailliert zu untersuchen. Von großer Bedeutung für die Umsetzung der Arbeit war die durchgeführte Entwicklung und Integration eines automatisierten Datenanalyseschemas. Besonders hervorzuheben sind die kontinuierliche Charakterisierung der Luftmassenherkunft, die Auswertung von multiplen Maxima im Wolkenradar-Dopplerspektrum, eine Methode zur Erkennung von durch Schwerewellen beeinflussten Wolken mit Doppler Lidar und die Integration aller Datenquellen in die verteilte LACROS-Forschungsdatenanwendung. Wichtigste Ergebnisse dieser Arbeit sind, dass atmosphärische Schwerewellen die Bildung und Detektierbarkeit der Eisphase erschweren und dass eine Kopplung von Wolken mit der planetaren Grenzschicht die Häufigkeit der Eisbildung erhöht. Wenn diese beiden Effekte berücksichtigt werden, tritt Eisbildung in Schichtwolken über Punta Arenas etwas weniger häufig auf als über Limassol und Leipzig. Dieser Unterschied kann auf eine geringere Verfügbarkeit von Eiskeimen in der freien Troposphäre über Punta Arenas zurückgeführt werden.:1 Introduction 2 Heterogeneous ice formation in shallow mixed-phase clouds 3 The mobile ground-based remote-sensing facility LACROS 3.1 LACROS instruments 3.1.1 MIRA-35 cloud radar 3.1.2 PollyXT multi-wavelength lidar 3.1.3 StreamLine XR Doppler lidar 3.1.4 Additional instruments and auxillary datasets 3.2 Campaigns under study 3.2.1 CyCARE field campaign 3.2.2 DACAPO-PESO field campaign 3.2.3 Observations at Leipzig 4 Methods and advancements in data processing 4.1 LACROS Research Data Application 4.2 Aerosol statistics based on the PollyNET processing chain 4.3 Estimating moments from radar Doppler spectra 4.4 Synergistic retrieval Cloudnet 4.5 Automated cloud identification 4.6 Gravity-wave detection 4.7 Continuous airmass source attribution 4.8 Transforming the Doppler spectrum into a tree structure 5 Contrasts in temperature, cloud and aerosol profiles 5.1 Occurrence of heterogeneous freezing regime 5.2 Cloud frequency 5.3 Profiles of airmass source 5.4 Aerosol optical properties 5.5 Lidar-based estimate of INP profiles 6 Properties of supercooled stratiform clouds 6.1 Overview on observed clouds 6.2 Case studies 6.2.1 Punta Arenas, 4/5 September 2019: stratiform cloud with variable ice formation 6.2.2 Punta Arenas, 27 September 2019: Wave cloud 6.2.3 Punta Arenas, 12 June 2019: Surface coupling 6.3 Phase occurrence frequency 6.4 Context to lidar-only observations 6.5 Effect of boundary-layer aerosol load on phase occurrence 6.6 Gravity-wave influence on phase occurrence at low temperatures 6.7 Ice-formation frequency of free-tropospheric and fully turbulent clouds 6.8 Contrasts of radar reflectivity factor in the ice virga 7 Multi-peak occurrence statistics of deeper clouds 8 Summary, Conclusions, and Outlook 8.1 Summary and conclusions 8.2 Outlook A Further equations Publication record List of Abbreviations and Acronyms List of Symbols BibliographyThis work investigates hemispheric contrasts of ice formation in stratiform supercooled liquid clouds using observations of three long-term campaigns of the mobile ground-based remote-sensing supersite LACROS (Leipzig Aerosol and Cloud Remote Observations System). For the northern hemisphere, LACROS datasets collected at Leipzig (Germany, 51.4°N, 12.4°E) and Limassol (Cyprus, 34.7°N, 33.0°E) between 2014 and 2018 were used. A key component of this work was the implementation of the more than two-year-long deployment of LACROS as part of the Dynamics Aerosol Clouds And Precipitation Observation in the Pristine Environment of the Southern Ocean (DACAPO-PESO) field campaign at Punta Arenas (Chile, 53.1°S, 70.9°W). The dataset assembled during this campaign resembles the first comprehensive multi-year ground-based remote-sensing dataset in the western part of the Southern Ocean. The synergistic combination of radar and lidar, including the capability to observe vertical velocities, allows detailed investigation of aerosol-cloud-dynamics interaction. One major part of this work was the development and integration of an automated data analysis scheme. Highlights are a continuous time-height-resolved airmass source characterization, a multi-peak analysis algorithm for radar Doppler spectra, a gravity-wave identification method based on Doppler lidar-vertical velocity observation, and the integration of the data sources into the distributed LACROS Research Data Application. The most important results of this work were that atmospheric gravity waves impede the formation and detectability of the ice phase, whereas the coupling of clouds with the planetary boundary layer increases the frequency of ice formation. When these two effects are taken into account, ice formation in stratiform clouds over Punta Arenas occurs slightly less frequent than over Limassol and Leipzig. This difference can be attributed to a lower availability of ice nuclei in the free troposphere over Punta Arenas.:1 Introduction 2 Heterogeneous ice formation in shallow mixed-phase clouds 3 The mobile ground-based remote-sensing facility LACROS 3.1 LACROS instruments 3.1.1 MIRA-35 cloud radar 3.1.2 PollyXT multi-wavelength lidar 3.1.3 StreamLine XR Doppler lidar 3.1.4 Additional instruments and auxillary datasets 3.2 Campaigns under study 3.2.1 CyCARE field campaign 3.2.2 DACAPO-PESO field campaign 3.2.3 Observations at Leipzig 4 Methods and advancements in data processing 4.1 LACROS Research Data Application 4.2 Aerosol statistics based on the PollyNET processing chain 4.3 Estimating moments from radar Doppler spectra 4.4 Synergistic retrieval Cloudnet 4.5 Automated cloud identification 4.6 Gravity-wave detection 4.7 Continuous airmass source attribution 4.8 Transforming the Doppler spectrum into a tree structure 5 Contrasts in temperature, cloud and aerosol profiles 5.1 Occurrence of heterogeneous freezing regime 5.2 Cloud frequency 5.3 Profiles of airmass source 5.4 Aerosol optical properties 5.5 Lidar-based estimate of INP profiles 6 Properties of supercooled stratiform clouds 6.1 Overview on observed clouds 6.2 Case studies 6.2.1 Punta Arenas, 4/5 September 2019: stratiform cloud with variable ice formation 6.2.2 Punta Arenas, 27 September 2019: Wave cloud 6.2.3 Punta Arenas, 12 June 2019: Surface coupling 6.3 Phase occurrence frequency 6.4 Context to lidar-only observations 6.5 Effect of boundary-layer aerosol load on phase occurrence 6.6 Gravity-wave influence on phase occurrence at low temperatures 6.7 Ice-formation frequency of free-tropospheric and fully turbulent clouds 6.8 Contrasts of radar reflectivity factor in the ice virga 7 Multi-peak occurrence statistics of deeper clouds 8 Summary, Conclusions, and Outlook 8.1 Summary and conclusions 8.2 Outlook A Further equations Publication record List of Abbreviations and Acronyms List of Symbols Bibliograph

Wave modelling in coastal and inner seas

In the long term development of the research on wind waves and their modelling, in particular of the inner and coastal seas, the present situation is framed with a short look at the past, a critical analysis of the present capabilities and a foresight of where the field is likely to go. After a short introduction, Chapter 2 deals with the basic processes at work and their modelling aspects. Chapter 3 highlights the interaction with wind and currents. Chapter 4 stresses the need for a more complete, spectral, approach in data assimilation. Chapter 5 summarizes the situation with a discussion on the present status in wave modelling and a look at what we can expect in the future.Peer ReviewedPostprint (author's final draft