VictorJuiz/thesis huggingface dataset:
Discovery of FeFe hydrogenase variants with enhanced O2 tolerance [electronic resource]

Hydrogenases catalyze the reversible formation of H2 from two protons and two electrons. They are classified as [FeFe], [NiFe], or [Fe]-only with respect to the metal atoms included in the active site. Among the three, [FeFe] hydrogenases have a catalytic bias toward H2 formation. The most prolific enzymes require overpotentials lower than that for platinum, one of the best metal catalysts reported to date. Consequently, [FeFe] hydrogenases offer significant potential for biological production of H2. We have approached such applications with a particular focus on the [FeFe] hydrogenase from Clostridium pasteurianum (CpI). It is a 63.8 kDa protein with one of the highest reported H2 production-specific activities in the order of 1,500 s-1 (or an order of magnitude higher when coupled with an electrode). However, it is also a complex enzyme with three accessory [4Fe-4S] clusters and one [2Fe-2S] cluster that deliver electrons to or from the active site consisting of an [FeFe] sub-cluster bridged to a [4Fe-4S] cluster by a cysteinyl thiol. Much work has been done during the past decade to enable the heterologous expression, maturation, and purification of the active form of CpI in Escherichia coli. Based on these advances, we are now working on two distinct routes of biological H2 production using this enzyme: Fermentative and photosynthetic. Unfortunately, [FeFe] hydrogenases are highly sensitive to O2, imposing a significant technical barrier for biological H2 production. The half-life of CpI in air-saturated buffer is estimated to be only 2--3 min. We worked on three projects to overcome this limitation with the end goal of engineering more O2-tolerant CpI variants. We first developed physiological assays for characterizing the O2 sensitivity of [FeFe] hydrogenases. Next, we developed a highthroughput screening platform for measuring H2 production rates in a biochemical context, including these assays. Empowered by the results of these projects, we evaluated over 300 CpI mutants and discovered several mutations that significantly impact the O2 tolerance of the enzyme. The best combination of mutations enabled greater than 4-fold improvement in aerobic (5.0 vol% O2) H2 yield over an hour. Our work provide the tools that allow convenient assessment of the O2 tolerance of [FeFe] hydrogenases and engineered mutants. The throughput has been improved by more than 10-fold. More importantly, we showed the first successful case of engineering [FeFe] hydrogenases with higher O2 tolerance, further promoting the research towards biological production of H2 for use as a renewable fuel and chemical.

Keywords: Chemical Engineering.
Advisor: Swartz James R. primary advisor. Thesis advisor
Cochran Jennifer R. advisor. Thesis advisor
Dunn Alexander Robert advisor. Thesis advisor
Disk resonator gyroscope within a wafer-scale encapsulation process [electronic resource]

Recently, MEMS gyroscopes have been widely adopted into consumer devices thanks to their small size, low cost, and efficient power consumption. In particular, wine- glass mode MEMS gyroscopes have received attention because of their potential to enter into high-performance applications such as gyro-compassing, spacecraft navigation, and dead-reckoning. In this dissertation, we present the development of Disk Resonator Gyroscopes (DRG) within a wafer-scale encapsulation process. First, we describe the fundamental principles of wineglass mode MEMS gyro- scopes, design of the DRG, its fabrication platform, and the different material choices for the device. The wafer-scale encapsulation used in this work, known as epi-seal, has been proven to be a highly stable, robust MEMS fabrication process. Disk-shaped resonator design implemented in epi-seal shows a high quality factor (Q) in excess of 100,000 at the resonant frequency of 250kHz; this high Q, along with the ability to minimize frequency splits via different material choices, drastically improves the scale factor of the DRG. Next, we discuss the implementation of the rate measurement demodulation technique with the DRG. The rate measurement reveals an angle random walk (ARW) of 0.1 ̊/√hr and a bias instability of ~1 ̊/hr. These noise specifications are compatible with low-end tactical grade gyroscopes, which are currently dominated by fiber-optics gyroscopes (FOG) and ring-laser gyroscopes (RLG). In addition, parametric amplification effects are discussed in further improving the performance of the DRG. Furthermore, we demonstrate improvements in stability of the scale factor and bias output with single-chip ovenization (micro-oven) of a fully-encapsulated DRG. During temperature-controlled operation, the scale factor stability holds to less than 0.3% and the bias remains less than 0.2 ̊/s, allowing for stable operation over a wide operating temperature range (0-80 ̊C).

Keywords: Mechanical Engineering.
Advisor: Kenny Thomas William primary advisor. Thesis advisor
Howe Roger Thomas advisor. Thesis advisor
Tang Sindy (Sindy K.Y.) advisor. Thesis advisor
Dislocation dynamics of face-centered cubic metals and alloys [electronic resource]

Face-centered cubic (FCC) metals and alloys—such as aluminum, copper, and austenitic stainless steel—are ubiquitous in the automotive, aerospace, and oil and gas industries. To further our understanding of the nature of plastic deformation in these materials, we have utilized dislocation dynamics (DD) simulations. We begin with a study of time integration in dislocation dynamics, examining the time-step-limiting aspects of DD, and developing a new subcycling-based time integrator that improves efficiency 100-fold. We then utilize this time integrator to study the basics of plasticity in pure, single crystalline copper. DD simulations were run over a wide range of strain rates and initial dislocation densities, examining how the yield strength and hardening rate vary, and making comparisons against the available experimental data. A detailed study on the contribution of binary dislocation junctions to hardening is then presented, showing that these junctions are an essential ingredient for hardening to occur. We then go on to study the most common strengthening mechanisms employed in FCC metals: solid solution strengthening and precipitation strengthening. After deriving the suitable chemical potential for the solute atoms comprising a solid solution, the influence of an "atmosphere" of solutes surrounding a dislocation on the stress field and line tension of the dislocation are examined, showing that these effects are generally small. The drag force exerted on dislocations by their atmospheres is then studied. We find these drag forces are often larger than those due to lattice friction, and can influence plasticity in FCC materials significantly. To study precipitation strengthening, a new algorithm for simulating dislocation-precipitate interactions is developed, which allows for ellipsoidal inclusions with arbitrary aspect ratio and arbitrary misfit. The new formulation is used to study Orowan looping in overaged aluminum-copper alloys, which have plate-like precipitates. The results and methods presented here constitute a broad set of advancements towards a more sound understanding of plastic deformation in FCC metals and alloys.

Keywords: Mechanical Engineering.
Advisor: Cai Wei
Balch Dorian K.
Lew Adrian advisor. Thesis advisor
Nix William D. advisor. Thesis advisor
Does Calcineurin Regulate the Osmotic Stress Response in S. cerevisiae Through the Smp1 Transcription Factor?

Calcineurin is a conserved Ca2+/calmodulin-regulated phosphatase that regulates the T cell immune response, as well as many other processes, and is the direct target of immunosuppressant drugs (cyclosporin A and FK506). Because of its ubiquitous expression and prevalence in a broad spectrum of disorders, long term suppression of calcineurin also results in harmful side effects (Safa, Riella, and Chandraker 2013). The lack of sequence similarity in bona fide calcineurin substrates, save for conserved calcineurin docking sites ('PxIxIT' and 'LxVP' ), has hindered candidate substrate identification. However, there have been recent efforts in systematically screening potential substrates for these docking sites. This study investigates the possible interaction between calcineurin and Smp1, a yeast transcription factor that is activated via phosphorylation by Hog1 MAP Kinase (Hog1 MAPK) under hyperosmotic conditions. Using computational analysis, all possible 6-mers in Smp1 were screened for the potential to interact with calcineurin as a ‘PxIxIT’ site, generating seven high-scoring peptide sequences. The goal of this project was to investigate the potential interaction between Smp1 and calcineurin, using two approaches: 1) testing for direct binding of Smp1 and calcineurin in vitro, and 2) studying regulation of Smp1 activity by calcineurin in the presence of osmotic stress in vivo. Co-purification assays revealed that full length GST-tagged Smp1 binds to yeast calcineurin in vitro, and may bind at one of the computationally predicted ‘PxIxIT’ sites. In vivo functional assays revealed that calcineurin both positively and negatively regulated Smp1 activity, suggesting that calcineurin may regulate the Hog1 MAPK pathway through multiple substrates. Overall, these results suggest that calcineurin is intimately involved in the Hog1 MAPK pathway, and substantiates this computational approach for de novo identification of calcineurin interaction sites. This approach may be used for future identification of novel calcineurin substrates, and provide insight on how to selectively target calcineurin substrates.

Keywords:  Department of Biology
Biology
Calcineurin
Phosphatase
Osmotic Stress
Smp1
Hog1 MAP Kinase
Thesis
Advisor: Cyert Martha
Bergmann Dominique
The drift and minorization method for reversible Markov chains [electronic resource]

This thesis proves upper bounds on the convergence time to stationarity for reversible discrete time Markov chains on general state spaces. The method of proof is the well-established drift and minorization approach, which imposes a regenerative structure on the Markov chain according to a particular recipe. The resulting bounds are computable in terms of the ingredients of the recipe. The convergence theorems in this thesis are developed using a new perspective on the interplay between regeneration and reversibility. They are more widely applicable than previous results and provide better numerical bounds on the time to stationarity in specific examples. Two Gibbs samplers from Bayesian statistics are treated in detail. When applied to these chains, the new bounds improve on earlier work but are still quite conservative compared with the true convergence rates. For certain classes of finite chains, the drift and minorization method can give precise bounds on the mixing time. A striking result of this type is a sharp upper bound on the cutoff window for birth and death chains. In addition, an inductive argument shows that the spectral gap of the random walk on the hypercube can be recovered using drift and minorization up to a constant factor of 2. The thesis also contains: (1) An exposition of the drift and minorization method, explaining both the renewal theory approach of Meyn and Tweedie and the coupling approach of Rosenthal, and showing how the bounds improve when the chain is reversible or stochastically monotone. (2) A development of the properties of different types of regeneration times for general state space Markov chains. Defining a randomized stopping time usually requires enlarging the sample space to incorporate independent randomness. In full generality, this operation raises subtle questions of measurability, which are resolved using a new "compatibility condition." (3) A brief consideration of quantile estimation in Markov chain Monte Carlo, focusing on concentration inequalities that provide finite-sample guarantees at specified confidence levels. The goal is to prove inequalities that rely as little as possible on theoretical convergence bounds (of the sort proved elsewhere in this thesis) and as much as possible on the empirical sample.

Keywords: Mathematics.
Advisor: Diaconis Persi primary advisor. Thesis advisor
Chatterjee Sourav advisor. Thesis advisor
Zheng Tianyi
Dual tumor suppressive and pro-oncogenic roles for the Notch signaling pathway in small cell lung cancer [electronic resource]

Developmental pathways are frequently re-activated and implicated during tumorigenesis. During lung development, Notch signaling inhibits the formation of pulmonary neuroendocrine cells, which are the adult epithelial cells of origin for small cell lung cancer (SCLC), the most aggressive and lethal subtype of lung cancer. This inhibitory role of Notch in neuroendocrine differentiation suggested that Notch activity might be tumor suppressive in SCLC. Consistent with this, genomic studies revealed inactivating mutations in the NOTCH receptors in ~15-25% of SCLC tumors. Ectopic Notch activation also inhibited tumor initiation in a mouse model for SCLC, suppressed the proliferation of neuroendocrine SCLC cells in culture and induced a differentiation switch to a non-neuroendocrine cell fate. Surprisingly, however, we identified a subset (~25%) of cells (Notch+ cells) in SCLC tumors that activate endogenous Notch signaling. These cells are non-neuroendocrine, proliferate slower than the Notch-, neuroendocrine tumor cells and are generated from the neuroendocrine cells, thus establishing a level of intratumoral heterogeneity generated by the bulk tumor cells themselves. Notch+ cells promote the proliferation of neuroendocrine cells, and Notch inhibition cooperates with chemotherapy to inhibit SCLC growth. These data show that the Notch pathway can be both tumor suppressive and oncogenic in SCLC; Notch inhibits the proliferation and differentiation of neuroendocrine cells but also promotes their growth in a paracrine manner. Downstream of Notch, we identified Rest/Nrsf, a transcriptional repressor that typically acts to restrict the expression of neuronal genes, as a target of Notch that mediates its inhibition of neuroendocrine cell fate. Thus, the Notch pathway has complex roles in SCLC and is able to modulate both tumor growth as well as its neuroendocrine differentiation.

Advisor: Sage Julien primary advisor. Thesis advisor
Krasnow Mark
Rando Thomas A.
Sweet-Cordero Eric advisor. Thesis advisor
Dynamic strategy for personalized medicine [electronic resource] : an application to metastatic breast cancer

Planning for treatment of cancer is challenging due to the complexity of the disease. Oncologists have to select and switch therapies considering the trade-off between treatment efficacy and therapy sides effects for an increasing number of therapies and combinations. Moreover, cancer varies with different patient characteristics and past treatment history. We develop a framework for computing a dynamic strategy for therapy choice in a large class of breast cancer patients, as an example of approaches to personalize therapies for individual characteristics and each patient's response to therapy. Our model maintains a Markov belief about the effectiveness of the different therapies and updates it as therapies are administered and tumor images are observed. We compare three different approximate methods to solve our analytical model against standard medical practice and show significant potential benefit of the computed dynamic strategies to limit tumor growth and to reduce the number of time periods patients are given chemotherapy, with its attendant side effects. We test robustness of our model with sensitivity analysis on key model parameters, seeing how the optimal dynamic strategy changes when patient characteristics differ. We also demonstrate the scalability of our model and recommended algorithm, showing that it has potential of providing real time advice to patients and oncologists.

Keywords: Management Science and Engineering.
Advisor: Shachter Ross D. primary advisor. Thesis advisor
Howard Ronald A. (Ronald Arthur)
Rubin Daniel
Dynamics of photosynthetic pigment-protein complexes in Chlamydomonas reinhardtii [electronic resource]

Photosynthesis, the conversion of solar energy into fixed chemical energy, is a critical biological process that supports nearly all life on earth. This process involves the absorption of light by pigment-protein complexes that make up thylakoid or photosynthetic membranes, and electron transfer reactions that ultimately generate NADPH and establish a proton gradient used to synthesize ATP. These cellular reducing equivalents and high energy compounds are then utilized in carbon fixation reactions that occur in the Calvin Benson Bassham Cycle (reductive pentose phosphate pathway). However, photosynthesis is also an inherently dangerous process as excess absorbed light that cannot be used for photochemistry (electron transport and CO2 fixation) can elicit the formation of reactive oxygen species that can severely damage the cell. Therefore, photosynthesis must be a highly regulated and extremely flexible process. We are just beginning to understand all of the components of the photosynthetic apparatus and how they function in the face of an ever-changing environment. This thesis explores novel components and functions associated with photosynthesis, with an emphasis on pigment-protein complex dynamics. All of the work was performed using the single-celled green alga Chlamydomonas reinhardtii, an excellent model for the study of photosynthesis. The first introductory chapter highlights unifying themes related to the control and plasticity of photosynthesis. The second chapter focuses on two novel thylakoid membrane proteins, CPLD38 and CPLD49, both of which are critical for photoautotrophic growth and impact the accumulation of the cytochrome b6f complex. The third chapter highlights a critical, yet previously undiscovered role for linear tetrapyrroles (bilins) in the maintenance of photosystem I. Central to this study is the control of photosynthetic processes as cells transition from darkness to light. The fourth chapter focuses on a critical redox carrier protein, FDX5, which is absolutely required for growth in the dark where it may be involved in maintaining proper membrane structure and composition, and other processes that are potentially redox-associated. The fifth chapter addresses how photosynthesis is adjusted during nutrient deprivation and specifically elucidates unique aspects of photoautotrophic nitrogen deprivation (an often encountered condition in the environment) that allow the photosynthetic apparatus to be preserved even though the cells are unable to fix much inorganic carbon. The sixth chapter presents preliminary evidence that identifies specific steps in the assembly of the photosynthetic apparatus that appear to be protected from atmospheric O2. Overall, this work unmasks novel components associated with the function and biogenesis of thylakoid membranes, revealing mechanisms critical for maintaining photosynthetic function in a dynamic environment.

Keywords: Biology.
Advisor: Grossman Arthur (Arthur R.) primary advisor. Thesis advisor
Mudgett Mary Beth
Cyert Martha S.
Spormann Alfred M. advisor. Thesis advisor
Dynamics of plasma discharges used for space propulsion [electronic resource]

Hall thrusters are a mature form of electric propulsion for spacecraft well suited for north-south station keeping, orbit transfer maneuvers, and eventually deep space travel. Despite decades of development, producing propulsion systems with high operating efficiency and specific impulse, key aspects of the physics governing these complex devices remain poorly understood and fundamental experimental and computational studies persist. This work concerns the development and implementation of laser-induced fluorescence (LIF) diagnostics for probing time-resolved dynamics in systems with a quasi-periodic forced or naturally occurring oscillation. Relaxing the requirement of a fixed frequency spectrum over time distinguishes the sample-hold and fast switching LIF methods studied here from others currently in use. The methods are validated against a collisional-radiative model of a 60 Hz xenon test discharge, successfully predicting the time evolution of excited xenon state populations. The sample-hold method is then applied to studying xenon neutral and ion dynamics and transport in two Hall thruster discharges: a laboratory model in the Stanford Plasma Physics Laboratory similar to the SPT-100 class of devices, and a commercial Busek Co. BHT-600 Hall thruster at the Air Force Research Laboratory at Edwards Air Force Base, California. Time-resolved ion velocity measurements in the channels and near-field plumes of these thrusters reveal propagating propellant ionization and acceleration fronts undergoing periodic motion that are fundamentally linked to the breathing mode ionization instability common to such devices. The presence of low ion velocity populations exhibiting different dynamic behavior than the main accelerated population is likely due to residual ionization of neutral propellant in the near-field plume of the thrusters. A study of the poorly understood central jet in the BHT-600 also reveals new details about the asymmetric nature of this plasma feature in both space and time. Finally, the measured ion dynamics in the Stanford Z-70 thruster are compared with those predicted by a two-dimensional hybrid particle-in-cell simulation of the device with a dynamically updating electron mobility modeled after the turbulent dissipation of energy in the electron fluid. Somewhat similar modulations in ion velocity are observed in the simulation results, but the limits of these types of simulations in tracking accurate time-resolved behavior are revealed.

Keywords: Mechanical Engineering.
Advisor: Cappelli Mark A. (Mark Antony) primary advisor. Thesis advisor
Hanson Ronald advisor. Thesis advisor
Hargus William A. advisor. Thesis advisor
Economic recovery and the determinants of productivity and innovation [electronic resource] : evidence from post-WWII Italy

This dissertation explores the process of economic development of Italy which between the late 1940s and the early 1970s moved from being a war-ravaged agricultural country to the seventh most industrialized power of the world. Chapter 1 investigates the extent to which the Marshall Plan affected the Italian economic recovery from WWII and its subsequent industrial production expansion. Chapter 2 assesses the long-run effects of management and technology transfer on firm performance. Chapter 3 studies the series of Italian reforms that promoted university STEM education among a group of high-school students, who were trained in STEM, but historically denied access to scientific university studies.

Keywords: Economics.
Advisor: Abramitzky Ran primary advisor. Thesis advisor
Bloom Nick
Dupas Pascaline advisor. Thesis advisor
Morten Melanie advisor. Thesis advisor