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Title: Aspects of Computational Topology and Mathematical Virology
Creator: Wang, Rui
Date: 2022
Collection: Electronic Theses & Dissertations
Description: Being able to describe the shape of data is of paramount importance to the fields of biology, physics, chemistry, pharmaceutics, etc. Therefore, in recent years, scientists from the TDA community have been applying advanced mathematical tools to decode the topological structures of data. Methods such as persistent homology, path homology, and de Rham-Hodge theory have become the main workhorse of TDA, which pioneered new branches in algebraic topology and differential geometry. Later, various...
Show moreBeing able to describe the shape of data is of paramount importance to the fields of biology, physics, chemistry, pharmaceutics, etc. Therefore, in recent years, scientists from the TDA community have been applying advanced mathematical tools to decode the topological structures of data. Methods such as persistent homology, path homology, and de Rham-Hodge theory have become the main workhorse of TDA, which pioneered new branches in algebraic topology and differential geometry. Later, various topological Laplacians such as graph Laplacian, Hodge Laplacian, sheaf Laplacian, and Dirac Laplacian are proposed to preserve topological invariants and geometric shapes simultaneously. However, such Laplacians fail to extract the topological and geometric deformations when one introduces the filtration parameters in. Therefore, we proposed a new topological Laplacians called persistent Laplacians to fully recover the topological persistence and homotopic shape evolution during filtration. It is worth mentioning that persistent Laplacians are insensitive to asymmetry or directed relations, which limits their power to preserve the directional information of structures in practical applications. Therefore, we proposed persistent path Laplacians to overcome this issue. Similar to the persistent Laplacians, one can also extract the topological persistence and geometric deformations during filtration from the persistent path Laplacians by calculating their harmonic and non-harmonic spectra. In addition, the persistent path Laplacians are constructed on the directed graphs or network, which address the importance of directional representation in datasets such as gene regulation datasets in biology. Versatile mathematical tools have been playing an essential role in various biological applications. Since the first COVID-19 case was reported in December 2019, researchers worldwide have been pursuing scientific endeavors in the SARS-CoV-2 projects. Instead of designing promising vaccines and antibody therapies that required wet lab resources, we proposed a new mathematical-AI model called TopNetmAb to systematically analyze the mutation-induced impacts on the SARS-CoV-2 infectivity, vaccines, and antibody drugs. In this dissertation, the topological data analysis (including the persistent Laplacians mentioned above), artificial intelligence, various network models, and genomics analysis are all included in our SARS-CoV-2-related projects to provide comprehensive representations for the understanding of the transmission and evolution of SARS-CoV-2.
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Title: Laboring for the motherland : A mother-artist-researcher-teacher's reconfiguration of the state-sponsored transnational teacher
Creator: Gordon, Rebekah R.
Date: 2022
Collection: Electronic Theses & Dissertations
Description: This arts-based inquiry explores historical and contemporary configurations of state-sponsored transnational teachers, or those who teach overseas in programs that are financially and/or programmatically supported by their home government. In the U.S., some examples of state-sponsored transnational teaching programs include the Peace Corps, the Fulbright Specialist Program, and the English Language Fellow Program. Standing on feminist grounding, I draw upon maternal concepts from the field of...
Show moreThis arts-based inquiry explores historical and contemporary configurations of state-sponsored transnational teachers, or those who teach overseas in programs that are financially and/or programmatically supported by their home government. In the U.S., some examples of state-sponsored transnational teaching programs include the Peace Corps, the Fulbright Specialist Program, and the English Language Fellow Program. Standing on feminist grounding, I draw upon maternal concepts from the field of mother(hood) studies to frame my exploration and to interrogate and disrupt the patriarchal systems in which state-sponsored transnational teaching is situated. More specifically, I extend a/r/tographic methodology to include my identifications and embodied perspectives as a mother-artist-researcher-teacher. My proposed methodology of m/a/r/tography affords me the opportunity to think intergenerationally about the ways that orientations, collective body memory, family histories, and national legacies relevant to state-sponsored transnational teaching are passed down. By using epistolary narrative and creative non-fiction, I reflect on my own experiences as a mother, as a daughter, and as a former state-sponsored transnational teacher who labored for her motherland. To analyze these personal experiences, I adopt a diffractive approach in which these experiences are viewed through one another as well as through additional sources of data, including interviews with other state-sponsored transnational teachers, letters from my mother, photographs and images, poetry, film, children’s literature, and other forms of scholarship. The use of a diverse and wide-ranging set of scholarship is an attempt to not only spark creative connections between seemingly disparate sources of data but also inspire more accessible and humanizing ways of doing research and re-imagining (teacher) education that welcomes processes of re/dis/orientation.
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Title: LEAN IN : THE ANTECEDENTS AND CONSEQUENCES OF FEMALE DIRECTORS’ ATTAINMENT OF POWERFUL POSITIONS ON BOARDS
Creator: Kim, Jooyoung
Date: 2022
Collection: Electronic Theses & Dissertations
Description: With the heightened societal demand for promoting egalitarian values in corporate boards, firms have devoted much effort to improving female board representation. However, recent insights have suggested that some progress made thus far may be the result of firms’ symbolic conformity to external pressure, which alludes to the possibility that female directors’ low influence and lack of integration on boards may still persist. In this dissertation, I aim to develop and test theoretical...
Show moreWith the heightened societal demand for promoting egalitarian values in corporate boards, firms have devoted much effort to improving female board representation. However, recent insights have suggested that some progress made thus far may be the result of firms’ symbolic conformity to external pressure, which alludes to the possibility that female directors’ low influence and lack of integration on boards may still persist. In this dissertation, I aim to develop and test theoretical arguments of what determines and follows female directors’ attainment of power and influence, as reflected in their assignment to major committee member or chair positions. First, I propose that female directors are at a relative disadvantage in attaining major committee positions. Specifically, although directors in general can benefit from expertise cues to be assigned to committee positions, the benefits are less for females than males. This gap can be attenuated when females are similar to board members in terms of other demographic dimensions and when the board is demographically heterogeneous. Second, I develop predictions about how female representation on board committee positions can contribute to gender diversity on boards and firm performance. I propose that greater female representation on board committees has negative relationship with a female director’s likelihood of exit, and has positive relationship with female additions to the board and firm performance. Results were drawn from a sample of S&P 1,500 companies during 2009 to 2019.
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Title: Role of circular exercise on forelimb loading and accompanying skeletal and joint adaptations
Creator: Logan, Alyssa A.
Date: 2022
Collection: Electronic Theses & Dissertations
Description: Circular exercise is used frequently in equine exercise and competition, but little is known of the impact circular diameter and gait have to the joint and bone health of the forelimb. The first study evaluated the impact of circle diameter (10-m and 15-m) and gait to the forelimb solar outputs of average surface area, vertical force, and average pressure. Nine horses exercised in a straight line and in a round pen while wearing the Tekscan Hoof SystemTM on both front hooves with a glue-on...
Show moreCircular exercise is used frequently in equine exercise and competition, but little is known of the impact circular diameter and gait have to the joint and bone health of the forelimb. The first study evaluated the impact of circle diameter (10-m and 15-m) and gait to the forelimb solar outputs of average surface area, vertical force, and average pressure. Nine horses exercised in a straight line and in a round pen while wearing the Tekscan Hoof SystemTM on both front hooves with a glue-on shoe, a method of adherence which was determined to be reliable when measurements were recorded within one session. Gait, and not circle diameter, impacted forelimb outputs, with the average loaded area of the outside hoof while circling, being greatest at the canter (P = 0.001). While exercising on both a large and small circle, the outside hoof had greater vertical force at the canter than the trot (P = 0.01). A second study utilizing calves as a model for juvenile horses allowed the determination of physiological responses to circular exercise. Calves were assigned to small circle exercise (12 m), large circle exercise (18 m), treadmill exercise, or non-exercised control treatments (n = 6). Computed tomography and biomarkers were evaluated to determine impacts to bone and joint health. The inside leg of the small circular exercise group had larger dorsopalmar external diameter than the outside (P = 0.05). The medial proximal phalanx had greater mediolateral diameter than the lateral proximal phalanx of the small circle group (P = 0.01). Cartilage glycosaminoglycan concentration was greater in the outside leg of the small circle exercise treatment than the inside leg (P = 0.03). Combined, both of these studies suggest that circular exercise diameter and gait can impact animal health and should be considered when performing circular exercise.
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Title: Causal Inference with Mendelian Randomization for Longitudinal Data
Creator: Qu, Jialin
Date: 2022
Collection: Electronic Theses & Dissertations
Description: Mendelian Randomization (MR) uses genetic variants as instrumental variables (IVs) to examinethe causal relationship between an exposure and an outcome in observational studies. When confounding factors exist, the correlation between a predictor variable and an outcome variable does not imply causation. IV regression has been a popular method to control the confounding effect for causal inference. According to Mendel’s first and second laws of inheritance, genetic variants can be considered...
Show moreMendelian Randomization (MR) uses genetic variants as instrumental variables (IVs) to examinethe causal relationship between an exposure and an outcome in observational studies. When confounding factors exist, the correlation between a predictor variable and an outcome variable does not imply causation. IV regression has been a popular method to control the confounding effect for causal inference. According to Mendel’s first and second laws of inheritance, genetic variants can be considered as valid IVs. Popular MR methods include the ratio estimator, the inverse-variance weighted estimator and the two stage estimator. However, all these methods are based on cross-sectional data. In practice, data in the observational studies can be collected over time, the so-called longitudinal data. Longitudinal data makes it possible to capture changes within subjects over time and thus offers advantages to causal modeling to establish causal relationships. However, causal inference method that can control the time-varying confounding effect is largely lacking in literature. In this dissertation, we explore MR analysis for longitudinal data by proposing different causal models and assuming different casual mechanisms. The proposed methods are strongly motivated by a real study to examine the causal relationship between hormone secretion and emotional eating disorder in teen girls. We start with a concurrent model which assumes current outcome is only affected by current exposure. Coefficients of both genetic variants (i.e., IVs) and exposure are considered as time- varying effects. We apply the quadratic inference function approach in a two-step IV regression framework and focus on statistical testing to infer causality. Through extensive simulation studies, we show that the proposed method can well protect type I error and has reasonable testing power. In Chapter 3, we generalize the concurrent model to a more complex case and propose a time lag model to investigate time delayed causal effects. In the time lag model, we assume current outcome at time ? is affected by previous exposures measured up to ? − ? time points, where the time lag △? can be determined by a rigorous model selection procedure based on data. Similar to the concurrent model, we assume the effects of genetic variants on exposure and the effects of exposure on outcome both are time-varying. We propose different tests for point-wise and simultaneous testing to assess the causal relationship. In Chapter 4, We further generalize the time lag model to the case where the cumulative effect of previous ? exposures contributes to the outcome at time ?, under a sparse functional data analysis framework. The causal relationship is examined under the functional principal component regression framework with sparse functional data. Simulation results show that the type I error is well controlled. We apply our models to the emotional eating disorder data to examine if hormone secretion during the menstrual cycle in teen girls has a causal effect on emotional eating behavior and identify interesting results. This thesis work represents the very first exploration in MR analysis with longitudinal data.
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Title: PHENYLENEDIAMINE PYRIDYL LIGANDS AND BORYL SUPPORT LIGANDS FOR ORTHO-DIRECTED IRIDIUM CATALYZED C–H BORYLATION
Creator: O'Connell, Alex C.
Date: 2022
Collection: Electronic Theses & Dissertations
Description: With organoboron compounds being useful components in the synthesis of pharmaceuticals, agrochemicals, and materials, it is imperative to find new catalytic strategies to design an effective system capable of borylating a broad range of (hetero)arene substrates in high yields and high selectivity. Traditional iridium-catalyzed systems borylate aromatic compounds and are directed by steric factors of the substrate. These steric-directed catalysts are hypothesized to have a singly open...
Show moreWith organoboron compounds being useful components in the synthesis of pharmaceuticals, agrochemicals, and materials, it is imperative to find new catalytic strategies to design an effective system capable of borylating a broad range of (hetero)arene substrates in high yields and high selectivity. Traditional iridium-catalyzed systems borylate aromatic compounds and are directed by steric factors of the substrate. These steric-directed catalysts are hypothesized to have a singly open coordination site on the metal center where activation of the most accessible C–H bond can occur. In order to change regioselectivity from steric products to alternatives, new catalyst systems must be designed.A phenylenediamine pyridyl framework was implemented for chelate-directed C–H borylation, where an aromatic substrate undergoes borylation of the ortho C–H bond, relative to a directing group. This ligand type has been explored and shown to have three major components that influence the reactivity, selectivity, and coordination of the ligand. These parts that make up the ligand were examined using a ligand screen, NMR studies, and stoichiometric reactions. From the literature, it has been shown that double B,N-bidentate ligated catalysts work well for a broad substrate scope and produce borylated products whose substitution pattern is based on steric effects. Other variants of this system have used a single B,N-bidentate ligand to produce products borylated in the ortho-position relative to a directing group on the substrate. To improve upon these catalytic systems, experiments were performed to optimize ortho-selectivity of the originally steric-directed catalyst containing two B,N-bidentate ligands by reducing the loading of the dimer boryl ligand. In doing so, regioselectivities can be completely switched from steric products to chelate products. This modification of ligand to metal ratio greatly effects selectivity and is a unique feature to dimer boryl ligands. These phenylenediamine pyridyl ligands and boryl support ligands will be explored.
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Title: EMOTIONALLY FOCUSED THERAPY FOR COUPLES IN TAIWAN
Creator: Tseng, Chi-Fang
Date: 2022
Collection: Electronic Theses & Dissertations
Description: This is the first study to begin to explore the effectiveness and predictors of change of emotionally focused therapy for relationship satisfaction and depressive symptoms among couples in Taiwan. This one-arm pragmatic trial assessed the clinical outcomes of 17 couples using paired-samples t-tests and multilevel modeling. Paired-sample t-tests revealed no statistical differences in relationship satisfaction and depressive symptoms before and after EFT. Additionally, multilevel modeling...
Show moreThis is the first study to begin to explore the effectiveness and predictors of change of emotionally focused therapy for relationship satisfaction and depressive symptoms among couples in Taiwan. This one-arm pragmatic trial assessed the clinical outcomes of 17 couples using paired-samples t-tests and multilevel modeling. Paired-sample t-tests revealed no statistical differences in relationship satisfaction and depressive symptoms before and after EFT. Additionally, multilevel modeling indicated no change in relationship satisfaction over time. However, there was a quadratic change in depressive symptoms over the course of EFT. While the study results were unexpected, it is important to note that most couples did not receive the recommended treatment “dose” in this pragmatic trial. In addition, the small sample size limited statistical power. In addition to assessing treatment outcomes, predictors of change were also examined. Findings showed that high traditionalism at intake predicted an increase in relationship satisfaction for women. Attachment was also a significant predictor of change; men with high attachment avoidance at intake demonstrated a significant decrease in depression, and men and women with high attachment anxiety at intake also experienced a significant decrease in depression. Lastly, emotional expressivity at intake was associated with an increase in relationship satisfaction and a decrease in depression for both men and women. Our study suggested that traditionalism, attachment, and emotional expressivity are important predictors of change among couples in Taiwan who receive EFT. While more research is needed, these findings offer preliminary support for the types of partners who may be more likely to experience change after receiving EFT.
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Title: EFFECTS OF PLACENTAL LISTERIA MONOCYTOGENES INFECTION ON FETAL NEURODEVELOPMENT
Creator: Lee, Kun Ho
Date: 2022
Collection: Electronic Theses & Dissertations
Description: Maternal infection can lead to adverse pregnancy outcomes. Numerous epidemiological studies have demonstrated an association between prenatal infection and neuropsychiatric disorders, including autism spectrum disorder (ASD). Different prenatal infections are associated with distinct neurological pathologies, necessitating studies of the diversity of prenatal pathogens and their consequences. Listeria monocytogenes (Lm) is a foodborne pathogen that causes listeriosis, which typically affects...
Show moreMaternal infection can lead to adverse pregnancy outcomes. Numerous epidemiological studies have demonstrated an association between prenatal infection and neuropsychiatric disorders, including autism spectrum disorder (ASD). Different prenatal infections are associated with distinct neurological pathologies, necessitating studies of the diversity of prenatal pathogens and their consequences. Listeria monocytogenes (Lm) is a foodborne pathogen that causes listeriosis, which typically affects immunocompromised individuals, including pregnant mothers. Prenatal infection with Lm can cause detrimental pregnancy outcomes, such as miscarriages, stillbirths, preterm labor, and death in newborns. However, neurological outcomes of maternal listeriosis have not been characterized. Here, I sought to investigate whether placental infection with Lm is associated with altered neurodevelopment by using a bioluminescence strain of Lm and a murine model of pregnancy-associated listeriosis. I show that placental infection affects neurodevelopment during pregnancy and behavior in the offspring.To investigate how placental infection with Lm dysregulates fetal brain development, I performed RNA-seq on fetal brains to quantify the enrichment of genes that were associated with the infection during gestation. The findings of RNA-seq analysis illustrated that placental infection with Lm altered fetal brain transcriptome and showed sexually dichotomous gene expression profiles. I further assessed the effects of different traits, including Lm exposure, the intensity of placental infection, and sex on the fetal transcriptome using systems biology. The genes were grouped into co-expression modules. Notably, maternal infection and its intensity measured by bioluminescence imaging signal were significantly associated with specific modules, suggesting these traits are the main factors driving these transcriptional changes. Lastly, I showed that placental Listeria infection enriched ASD-associated genes. These results demonstrate that maternal listeriosis dysregulates fetal brain transcriptome during gestation. Neurodevelopment is a complex process influenced by various environmental factors during pregnancy. To examine whether prenatal infection with Lm affects cortical lamination and neural activity, I performed hematoxylin and eosin staining and immunohistochemistry. Gross anatomy of the brain structure analysis showed that placental infection with Lm affected cortical lamination in a localized manner. Furthermore, increased neural activity was observed in Lm- exposed male offspring. These results illustrate that placental infection with Lm induces morphological changes in brain tissue during neurodevelopment. Behavioral symptoms of neuropsychiatric disorders are an important component of the diagnosis. Animal behavioral assays and tools have been developed to examine animal behavior such as social interactions, anxiety, and repetitive behaviors. I examined behavior tests that resembled ASD to determine if mouse offspring born following placental infection displayed abnormal behavior. Lm-exposed offspring exhibited altered behaviors and showed sex-dependent behavioral changes. Overall, my work highlights the impact of maternal listeriosis on brain development during pregnancy and its effects on offspring’s behavior and contributes to the understanding of the spectrum of fetal neurodevelopment.
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Title: Multi-Modality Nondestructive Evaluation Techniques for Inspection of Plastic and Composite Pipeline Networks
Creator: Alzuhiri, Mohand
Date: 2022
Collection: Electronic Theses & Dissertations
Description: The extensive adoption of plastic pipelines is a growing phenomenon in different fields of the industry, with applications that extend from municipal water and sewer systems to the water lines in nuclear reactors. The large-scale adoption is motivated by the unique features of plastics like corrosion and chemical resistance, low cost, and design flexibility. While the dielectric nature of plastic pipelines provides unique design capabilities, it also introduces new challenges for the...
Show moreThe extensive adoption of plastic pipelines is a growing phenomenon in different fields of the industry, with applications that extend from municipal water and sewer systems to the water lines in nuclear reactors. The large-scale adoption is motivated by the unique features of plastics like corrosion and chemical resistance, low cost, and design flexibility. While the dielectric nature of plastic pipelines provides unique design capabilities, it also introduces new challenges for the operators when it comes to inspecting and ensuring the integrity of these pipelines’ networks. In this study, a multi-modal approach is adopted to address the threats affecting the safety of small diameter plastic pipelines and explore possible inspection solutions for emerging materials like composites. Structured light endoscopes with RGB-D inspection capability were developed for the inspection of surface defects in small diameter pipelines with novelties a) Design and miniaturization of RGB-D structured light sensor with electronic stabilization, b) Development of an algorithm to automatically calibrate the sensor when placed in a cylindrical environment, c) Design of a single shot phase measurement SL sensor that employs the sensor movement to improve the 3D reconstruction, and d) Design a stereoscopic SL sensor for 360-degree inspection. EM-based inspection was adopted to inspect subsurface defects and classify materials around the inspected pipelines. An investigative study was performed to test the probability of detecting cold fusion in butt fusion joints by using emerging NDE techniques, and a coplanar capacitive sensor was designed for the detection of legacy crossbores in gas pipelines. Finally, a thermoacoustic imaging system was developed in this study with potential applications for the inspection of composites and medical imaging. The novelties of this work can be summarized as follows: a) Development of a simulation model to study the thermoacoustic waves generation and the effect of multiple experimental parameters on the performance of thermoacoustic imaging systems, b) Improving the signal to noise ratio of pulsed TAI imaging systems by adoption non-coherent pulse compression. In summary, this study presents a multi-modal approach for the inspection of pipeline networks by adopting optical RGB-D imaging sensors for surface inspection, EM-based sensors for subsurface inspection and classification of objects outside the pipe, and finally, a hybrid imaging method with potential applications in medical imaging and inspection of composites.
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Title: SIGNALING MECHANISMS OF PULMONARY ARTERIAL HYPERTENSION
Creator: Ji, Yajing
Date: 2022
Collection: Electronic Theses & Dissertations
Description: Pulmonary arterial hypertension (PAH) is a severe and life-threatening disease that is characterized by elevated pulmonary blood pressure. A challenge in treating PAH is that while the current generation of therapeutics alleviate symptoms, they fail to target the underlying causes of the disease. Initially it was thought that PAH is caused by increased pulmonary vasoconstriction; it is now understood that PAH mainly results from remodeling of the pulmonary vasculature. Further...
Show morePulmonary arterial hypertension (PAH) is a severe and life-threatening disease that is characterized by elevated pulmonary blood pressure. A challenge in treating PAH is that while the current generation of therapeutics alleviate symptoms, they fail to target the underlying causes of the disease. Initially it was thought that PAH is caused by increased pulmonary vasoconstriction; it is now understood that PAH mainly results from remodeling of the pulmonary vasculature. Further characterization of the underlying mechanisms of PAH will identify newpharmacological targets to treat PAH. In this dissertation I seek to address this challenge from three distinct perspectives. In Chapter 2, I investigated the signaling network downstream of TGFβ and highlighted the MRTF/SRF pathway as potential therapeutical targets for PAH given its pivotal role regulating expression of contractile proteins in PASMCs. In Chapter 3, I aim to test whether TGFβ and the silencing of BMPR2, a member of the TGFβ family of receptors, contribute to the activation of lung fibroblasts in vitro. My results presented do not replicate the role of BMPR2 silencing found in other studies. This could be caused by the relatively short duration of BMPR2 silencing in our system. Finally, in Chapter 4, I perform a combined meta-analysis of several publicly available transcriptomic datasets of lung tissues from PAH patients. Using this approach, I identify PAH-associated signaling pathways, and chemical compounds which reverse a PAH-associated gene expression signature. My findings also suggest that while we bin PAH patients into various subtypes in the clinic, on a transcriptional level, PAH patients tend to group into distinct gene expression clusters without relying on their clinical subtype. These findings improve our understanding of PAH biology and also highlight several potential drug targets for PAH.
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Title: I. Determination of Absolute Configuration of Chiral 1,2-Diols. II. Progress Towards the Total Synthesis of Napyradiomycin A1.
Creator: Torabi Kohlbouni, Saeedeh
Date: 2022
Collection: Electronic Theses & Dissertations
Description: This dissertation focuses on two parts. The first part introduces an operationally simple and microscale method for the absolute stereochemical determination of 1,2-diols. In situ derivatization of 1,2-diols with dinaphthyl borinic acid generates the induced helicity of the two naphthyl groups, which leads to an observable ECCD spectrum. The observed P or M helicity follows a predictable trend for S and R chiral 1,2-diols, respectively.The Second chapter is the progress towards the asymmetric...
Show moreThis dissertation focuses on two parts. The first part introduces an operationally simple and microscale method for the absolute stereochemical determination of 1,2-diols. In situ derivatization of 1,2-diols with dinaphthyl borinic acid generates the induced helicity of the two naphthyl groups, which leads to an observable ECCD spectrum. The observed P or M helicity follows a predictable trend for S and R chiral 1,2-diols, respectively.The Second chapter is the progress towards the asymmetric catalytic synthesis of napyardiomycin A1. The chapter is divided to three sections. The first section is installation of chlorine chiral center at C3. This goal is achieved using cinchona chiral catalyst, and DCDMH as chloronium source. The second section is the synthesis of the -lapachone core of napyradiomycin A1, was accomplished using Diels-Alder/aromatization cascade reaction. The last section shows our effort toward the attachment of geranyl side chain.
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Title: Solving Computationally Expensive Problems Using Surrogate-Assisted Optimization : Methods and Applications
Creator: Blank, Julian
Date: 2022
Collection: Electronic Theses & Dissertations
Description: Optimization is omnipresent in many research areas and has become a critical component across industries. However, while researchers often focus on a theoretical analysis or convergence proof of an optimization algorithm, practitioners face various other challenges in real-world applications. This thesis focuses on one of the biggest challenges when applying optimization in practice: computational expense, often caused by the necessity of calling a third-party software package. To address the...
Show moreOptimization is omnipresent in many research areas and has become a critical component across industries. However, while researchers often focus on a theoretical analysis or convergence proof of an optimization algorithm, practitioners face various other challenges in real-world applications. This thesis focuses on one of the biggest challenges when applying optimization in practice: computational expense, often caused by the necessity of calling a third-party software package. To address the time-consuming evaluation, we propose a generalizable probabilistic surrogate-assisted framework that dynamically incorporates predictions of approximation models. Besides the framework's capability of handling multiple objectives and constraints simultaneously, the novelty is its applicability to all kinds of metaheuristics. Moreover, often multiple disciplines are involved in optimization, resulting in different types of software packages utilized for performance assessment. Therefore, the resulting optimization problem typically consists of multiple independently evaluable objectives and constraints with varying computational expenses. Besides providing a taxonomy describing different ways of independent evaluation calls, this thesis also proposes a methodology to handle inexpensive constraints with expensive objective functions and a more generic concept for any type of heterogeneously expensive optimization problem. Furthermore, two case studies of real-world optimization problems from the automobile industry are discussed, a blueprint for solving optimization problems in practice is provided, and a widely-used optimization framework focusing on multi-objective optimization (founded and maintained by the author of this thesis) is presented. Altogether, this thesis shall pave the way to solve (computationally expensive) real-world optimization more efficiently and bridge the gap between theory and practice.
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Title: FIBER-OPTIC SILICON FABRY-PEROT INTERFEROMETERS FOR HIGH-SPEED ANEMOMETER AND HIGH-SENSITIVITY BOLOMETER APPLICATIONS
Creator: Uddin, Nezam
Date: 2022
Collection: Electronic Theses & Dissertations
Description: Fiber-optic silicon Fabry-Perot interferometric temperature sensor offers the advantage of high-speed and high-resolution to characterize the ocean turbulence in oceanographic research. Compared to silica, the material that makes the optical fiber, silicon has a thermo-optic coefficient ten times higher and a thermal conductivity sixty time higher. Silicon is highly transparent in the infrared wavelength range and can be easily processed with the mature microfabrication technology. All of...
Show moreFiber-optic silicon Fabry-Perot interferometric temperature sensor offers the advantage of high-speed and high-resolution to characterize the ocean turbulence in oceanographic research. Compared to silica, the material that makes the optical fiber, silicon has a thermo-optic coefficient ten times higher and a thermal conductivity sixty time higher. Silicon is highly transparent in the infrared wavelength range and can be easily processed with the mature microfabrication technology. All of these make silicon a very attractive material for high-speed and high-resolution turbulence measurement. We attached a small silicon pillar to the end of an optical fiber to make fiber-optic Fabry-Perot interferometric sensor demodulated by a white light system for fast turbulence measurement. We studied the two modes of fiber-optic hot wire anemometer operation for turbulence measurement theoretically and experimentally. The constant temperature operation of the fiber-optic hot wire anemometer was introduced for the first time to reduce the time constant significantly. The anemometer used for demonstration is based on a silicon low-finesse Fabry-Perot interferometer (FPI) attached to the tip of a single mode fiber. Turbulent flow measurement method based on constant temperature operation offers high measuring speed, because the wire temperature is kept constant, the effect of thermal inertia of the wire is suppressed. We also investigated a new sensor structure experimentally and theoretically for the measurement of water flow with reduced directivity. This sensor consists of a laser heated silicon FPI embedded in a metal microsphere. Herein, the spherical shape of the outside metal shell gives a symmetric response to water flow direction; thus, the directivity is reduced greatly. Moreover, the water flow measurement by the hot wire fiber-optic water flow sensor based on laser heated silicon FPI need to compensate the effect of water temperature variation. We reported a technique to compensate the effect of water temperature change in the flow measurement by using another sensor which will track the temperature of the water. By using the information of the water temperature change, baseline can be defined which will provide unique wavelength change for the flow. Finally, the wavelength change corresponding to the flow speed were calibrated using the sensor pair after compensating the effect of water temperature variation. We expanded the use of silicon Fabry-Perot interferometric sensor in the measurement of plasma radiation by modifying the structure with gold coated silicon and multimode graded index fiber between the single mode fiber (SMF) and silicon. We reported the design, fabrication, and characterization of a fiber-optic bolometer (FOB) with improved noise equivalent power density (NEPD) performance and increased absorption to high energy photons by engineering the absorber of the FOB. We also have developed a multichannel fiberoptic bolometry system with five bolometers connected to each channel of the coarse wavelength division multiplexer (CWDM), a single light source of super luminescent LED (SLED) and a single I-MON 512 OEM spectrometer. Easy sensor fabrication, significantly enhanced measurement range compared to the previous high-finesse FPI bolometer system for measuring radiation are some of the advantages. Moreover, utilization of the FOB in the vacuum for radiation measurement with reduced time constant was also studied which is practically required in the fusion devices. This was done by adding a heat sink with the current FOB structure and using the deconvolution method to get better temporal resolution. Finally, the FOB with the heat sink was tested in the vacuum condition to measure the radiation using the deconvolution method. Experimental results are presented to support the idea of heat sink and deconvolution method for plasma radiation measurement.
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Title: KNOWLEDGE SPILLOVERS AND SAFE DRINKING WATER ACT COMPLIANCE
Creator: Redican, Kyle James
Date: 2022
Collection: Electronic Theses & Dissertations
Description: In the wake of the 2014 Flint Water Crisis, researchers, regulators, and utility professionals have given increased attention to understanding drivers of (CWS) Safe Drinking Water Act (SDWA) compliance by community water systems (CWSs). Most of this research has only explored system traits while ignoring the vital role of human capital, especially the operator. The status of CWS operators can vary widely between different systems. More critically, scholars have not investigated how effective...
Show moreIn the wake of the 2014 Flint Water Crisis, researchers, regulators, and utility professionals have given increased attention to understanding drivers of (CWS) Safe Drinking Water Act (SDWA) compliance by community water systems (CWSs). Most of this research has only explored system traits while ignoring the vital role of human capital, especially the operator. The status of CWS operators can vary widely between different systems. More critically, scholars have not investigated how effective external linkages between CWS operators have impacted SDWA compliance. Drawing from the theories of Organizational Learning’s inter-organizational learning, Innovation Systems’ knowledge transfers, and Agglomeration Economics’ knowledge spillovers, I hypothesized that increased interactions between CWS operators, facilitated in part by geographic proximity, would lead to more information sharing, increased CWS performance, and fewer SDWA violations. Remarkably little is known about the drivers of inter-operator interactions or whether such interactions improve SDWA compliance, and this research helped fill the data gap through a large-sample survey of CWS operators in Michigan to capture the frequency of interactions along with a range of operator and system characteristics which may explain why some operators participate in more inter-operator interactions than others. With this novel dataset, along with publicly available system and community data, this research first investigated what endogenous operator characteristics were associated with more reported inter-operator interactions. Through multiple methods on reported operator interactions, the Utility and Contract operators and operators with memberships in professional organizations appear more likely to report more interactions than Non-Affiliated operators and all operators who were not members of professional organizations. Second, based on Tobler’s first law of geography, there should be some spatial autocorrelation in the number of reported interactions, and this was tested using variogram modeling. Observed spatial autocorrelation indicated location-based differences in the number of reported interactions. Third, we used multiple methods to explore the primary research question to identify endogenous and spatial drivers of reported inter-operator interactions. Multiple models found that rural districts had a higher probability of fewer SDWA violations with increased interactions, while the urban districts had the inverse relationship. Fourth, the research incorporated CWS-specific and operator-specific variables, as the operator-specific data were not independent of the CWS observations (since some operators run multiple CWSs). I used a Generalized Linear Mixed-Model to estimate these relationships accounted for the multiple levels and found that more interactions increased the probability of SDWA compliance for certain types of operators. The broader implications of this research encourage stakeholders to pursue more inter-operator interactions as a low-cost mechanism to increase SDWA compliance. Seven avenues to increase interactions are outlined, ranging from open operator contact lists to operator focus groups to identify common problems and solutions to creating a state-level operator mentorship program to support new operators.
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Title: BUILDING STATE WILDLIFE AGENCY CAPACITY FOR EFFECTIVE PARTNERSHIPS
Creator: Cross, Megan M
Date: 2022
Collection: Electronic Theses & Dissertations
Description: State wildlife agencies (SWAs) partner with organizations of various types, on projects of various types, at what is anticipated to be an increasing rate. Inclusion of multiple and diverse stakeholders and partners is postulated to improve effectiveness of wildlife management (Anderson & Loomis, 2007; Jacobson et al., 2010). Through partnerships, actors from private, civil, and public sectors work together to reduce negative impacts from wildlife and improve access to and benefits of wildlife...
Show moreState wildlife agencies (SWAs) partner with organizations of various types, on projects of various types, at what is anticipated to be an increasing rate. Inclusion of multiple and diverse stakeholders and partners is postulated to improve effectiveness of wildlife management (Anderson & Loomis, 2007; Jacobson et al., 2010). Through partnerships, actors from private, civil, and public sectors work together to reduce negative impacts from wildlife and improve access to and benefits of wildlife resources. Although partnerships can improve the ability of SWAs to address these issues, little is known about how the perspectives of internal employees and external partners and stakeholders differ regarding factors affecting perceived success of partnerships in wildlife conservation.This dissertation addresses SWA partnerships through an examination of one prototypical SWA’s partnership arrangements. I propose a typology for categorization of SWA partnerships and apply a theory of collaborative capacity to the assessment of them. I surveyed all employees of the Michigan SWA and asked them to identify the three partners they consider most key to their work and found gaps in the frequencies of partners considered key to the work of SWA employees based on their locations in the defined typology. Additionally, the model of collaborative capacity tested varied in performance when applied to SWA employees and SWA partners. This research has implications for transparency regarding how state power is shared and considers how the disparate prevalence of various partnership arrangements may affect wildlife governance. Furthermore, my research findings may be used to improve SWA partnership arrangements and improve their alignment with governance and management-relate goals, as well as increase awareness of differences in views regarding partnership success as defined by SWA employees and partners.
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Title: High-precision and Personalized Wearable Sensing Systems for Healthcare Applications
Creator: Tu, Linlin
Date: 2022
Collection: Electronic Theses & Dissertations
Description: The cyber-physical system (CPS) has been discussed and studied extensively since 2010. It provides various solutions for monitoring the user's physical and psychological health states, enhancing the user's experience, and improving the lifestyle. A variety of mobile internet devices with built-in sensors, such as accelerators, cameras, PPG sensors, pressure sensors, and the microphone, can be leveraged to build mobile cyber-physical applications that collected sensing data from the real world...
Show moreThe cyber-physical system (CPS) has been discussed and studied extensively since 2010. It provides various solutions for monitoring the user's physical and psychological health states, enhancing the user's experience, and improving the lifestyle. A variety of mobile internet devices with built-in sensors, such as accelerators, cameras, PPG sensors, pressure sensors, and the microphone, can be leveraged to build mobile cyber-physical applications that collected sensing data from the real world, had data processed, communicated to the internet services and transformed into behavioral and physiological models. The detected results can be used as feedback to help the user understand his/her behavior, improve the lifestyle, or avoid danger. They can also be delivered to therapists to facilitate their diagnose. Designing CPS for health monitoring is challenging due to multiple factors. First of all, high estimation accuracy is necessary for health monitoring. However, some systems suffer irregular noise. For example, PPG sensors for cardiac health state monitoring are extremely vulnerable to motion noise. Second, to include human in the loop, health monitoring systems are required to be user-friendly. However, some systems involve cumbersome equipment for a long time of data collection, which is not feasible for daily monitoring. Most importantly, large-scale high-level health-related monitoring systems, such as the systems for human activity recognition, require high accuracy and communication efficiency. However, with users' raw data uploading to the server, centralized learning fails to protect users' private information and is communication-inefficient. The research introduced in this dissertation addressed the above three significant challenges in developing health-related monitoring systems. We build a lightweight system for accurate heart rate measurement during exercise, design a smart in-home breathing training system with bio-Feedback via virtual reality (VR) game, and propose federated learning via dynamic layer sharing for human activity recognition.
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Title: THE IMPACT OF ERAP1 FUNCTIONS ON INNATE AND ADAPTIVE IMMUNITY IN HUMAN DISEASE MODELS
Creator: Blake, Maja Kristin K.
Date: 2022
Collection: Electronic Theses & Dissertations
Description: ERAP1 has long been appreciated for its role in antigen presentation during the adaptive immune response. It is a peptidase in the endoplasmic reticulum that trims peptide antigens prior to their loading onto awaiting MHC-1. However, ERAP1 has also been shown to play an important role in innate immune responses, although the mechanisms underlying these associations have been unclear. The ERAP1 gene has also been linked to a number of autoimmune diseases including, but not limited to Multiple...
Show moreERAP1 has long been appreciated for its role in antigen presentation during the adaptive immune response. It is a peptidase in the endoplasmic reticulum that trims peptide antigens prior to their loading onto awaiting MHC-1. However, ERAP1 has also been shown to play an important role in innate immune responses, although the mechanisms underlying these associations have been unclear. The ERAP1 gene has also been linked to a number of autoimmune diseases including, but not limited to Multiple Sclerosis, Ankylosing Spondylitis, and Ulcerative Colitis. In addition, it is known to be altered in various tumor types as a means of immune evasion, and inhibitors for ERAP1 have shown promising results against tumors in vitro. Therefore, ERAP1’s significance in the susceptibility to diverse diseases is vast, and further study into how this protein participates in both innate and adaptive immune response mechanisms is justified. Our lab has previously published that immune cells and animals deficient in ERAP1 display proinflammatory phenotypes. In this dissertation, the mechanism as to how disruptions in normal ERAP1 function leads to proinflammatory phenotypes is studied. First, proinflammatory mechanisms within a critical innate immune cell, macrophages, are discerned using both ex vivo models and an in vivo inducible colitis mouse model. ERAP1 deficiency in the setting of a murine model of autoimmunity is also evaluated, revealing both disturbances in B cell development and function as dependent on normal ERAP1 activity, and that these disturbances can lead to exaggerated neuroinflammation in several murine models of MS. ERAP1 dependent proinflammatory mechanisms within B cells are further studied ex vivo using global RNA sequencing technology along with flow cytometry-based methods. Together, the results of these studies reveal that loss of ERAP1 function causes enhanced ER stress within the cell, leading to UPR activation, increased inflammasome activity, and evidence of increased pyroptosis. Given the broad spectrum of ERAP1 functions on immune cell functions, we capitalized on these insights to determine how ERAP1 inhibition might impact diseases such as cancer. Specifically, our results confirmed that that ERAP1 inhibition promoted NK cell directed tumor killing, a modality that had never been attempted until now. In conclusion, this dissertation capitalizes upon insights gained from human genetic studies associating ERAP1 with a variety of human disease susceptibilities, identifying the molecular mechanisms underlying these associations, and also illuminates possible new therapies for human diseases derived from study of ERAP1.
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Title: IDENTIFYING LEVERAGE POINTS TO CREATE RESILIENCE TO CLIMATE SHOCKS IN DRYLAND SOCIAL-ECOLOGICAL SYSTEMS
Creator: Sharma, Shubhechchha
Date: 2022
Collection: Electronic Theses & Dissertations
Description: Drylands cover approximately 40 percent of the earth’s land area and support more than a billion people, most of whom live in Sub-Saharan Africa (SSA), where agro-pastoralism is the major food system. Climate-related shocks such as drought, water-scarcity, diseases and pests, and food price spikes have profoundly impacted household food security among dryland agro-pastoralists, especially in Tanzania. Thus, there is a need to investigate mechanisms to ensure the future resilience of...
Show moreDrylands cover approximately 40 percent of the earth’s land area and support more than a billion people, most of whom live in Sub-Saharan Africa (SSA), where agro-pastoralism is the major food system. Climate-related shocks such as drought, water-scarcity, diseases and pests, and food price spikes have profoundly impacted household food security among dryland agro-pastoralists, especially in Tanzania. Thus, there is a need to investigate mechanisms to ensure the future resilience of livelihoods and food systems in these regions. The goal of this dissertation is to use resilience thinking approaches to identify points of intervention in dryland SES, to manage both present and future climate risks. Resilience assessment is guided by three major questions: “resilience of what, to what, and for whom”.Paper-I explores “resilience of what, to what, for whom” through the use of systems archetypes for a Maasai dryland agro-pastoralist food system in Northern Tanzania. The paper identified three system archetypes— Escalation, Limits to Growth, and Shifting the Burden—to (1) pinpoint the elements, patterns, and relationships that make up agro-pastoralist food systems; and (2) find leverage points to address the archetypical patterns limiting food security. The paper suggests a need for institutional strengthening and polycentricity to deal with food insecurity among agro-pastoralists. Paper-II explores what shocks Tanzanian food systems are responding to and how, i.e., “resilience of what, to what”. The paper used randomly sampled household data collected at national level through secondary sources to understand how different adaptive capacities influence their ability to deal with climate shocks, particularly with respect to ensuring food security, measured in terms of dietary diversity and household consumption expenditure. Through Structural Equation Modeling (SEM), the paper drew a path model that indicated investment in wealth and income diversification and investment in infrastructure were able to mediate the impact of shocks on food security. Paper-III similarly explores “resilience of what, to what” but using System Dynamics Modeling (SDM). The model allowed the exploration of feedback mechanisms and interactions between the population, livestock, and crop sub-sectors with food security in agro-pastoralist food systems in Naitolia village in Tanzania. The goal of the model was also to evaluate effectiveness of multiple policy scenarios required for food security. Out of four simulated scenarios, the model identified enhancing mechanisms for food production, along with reducing post-harvest losses and livestock predation, as most likely to result in a food sufficient scenario in the future. In creating these analyses and findings, the dissertation recommends four major leverage points to support food security through both present and future climate shocks : (1) maintaining diversity and redundancy in income and assets that provide insurance against failures; (2) fostering connectivity between multiple actors across networks for promoting bridging social capital; (3) ensuring polycentric governance so that the right well-connected institutions at the right time can deal with both agro-pastoralist rights to food and respond to disturbance and uncertainty. The dissertation also creates methodological advancement in the understanding of food security in complex systems under climate shocks, by utilizing a variety of approaches that support system thinking - systems archetypes, statistical modeling, and simulation through a system dynamics model.
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Title: When the War Raged On : Montana Territory, the Politics of Authority, and National Reconstruction
Creator: Andrella, Jennifer
Date: 2022
Collection: Electronic Theses & Dissertations
Description: In July 1861, the US House Committee on Territories drafted the first Reconstruction bill to detail a procedure for readmitting Southern states into the Union. Expecting a quick end to the Civil War, the earliest framers of Reconstruction recommended that rebellious states be assigned into an unorganized status as territories. It was a pragmatic solution that placed the South firmly in the control of a Republican Congress; a plan that complemented the Committee on Territories’ simultaneous...
Show moreIn July 1861, the US House Committee on Territories drafted the first Reconstruction bill to detail a procedure for readmitting Southern states into the Union. Expecting a quick end to the Civil War, the earliest framers of Reconstruction recommended that rebellious states be assigned into an unorganized status as territories. It was a pragmatic solution that placed the South firmly in the control of a Republican Congress; a plan that complemented the Committee on Territories’ simultaneous pursuit of territorial expansion in the trans-Mississippi West. Indeed, between 1861 and 1868 Congress incorporated seven Western territories to consolidate federal power in a growing domain. From the onset of the war, federal actors envisioned Reconstruction as a national process. Yet, the reality on the ground seldom matched their strategic plans. This dissertation analyzes Reconstruction from the vantage point of the Northwestern Great Plains. Using Montana Territory as a case study, I examine how relations between and among Native American nations, settlers, and government officials defined Reconstruction at both local and federal levels. The federal government had enduring political and economic interests in the Northwestern Plains prior to the outbreak of the war. Between 1828 and 1865, the region emerged as the last US stronghold of the global fur trade, cycled through several mining booms, and showed a promising future for homesteading and ranching. The Northwestern Plains were and are the homelands to a mosaic of Native American nations who asserted their rights to sovereignty by demanding federal recognition of their territorial, political, economic, and cultural autonomy. As these lands became contested under the pressure of US settlement, Native actors continued to press for visibility against local and federal modes of authority. The lived experiences of Native actors unveil some of the critical limitations of Reconstruction; that the expansion of citizenship, suffrage, and labor protections coincided with land dispossession, colonization, and erasure. By the time this study concludes in 1883, it becomes apparent that the dissolution of Reconstruction rested in the program’s failure to resolve the nation’s most fundamental questions over belonging, space, and power.I argue that Reconstruction was a process that experimented with federal and local forms of authority, settler colonialism, and state formation which came under stress after the onset of war in 1861. Republican governance throughout the Civil War and Reconstruction introduced new federal economic and political imperatives, destabilized local patterns of power among settlers, and opened new threats to Indigenous sovereignty. Using cartography, personal and mass communication, artwork, literature, and government records, this study portrays a version of Reconstruction that was fluid, chaotic, and often violent as western civil institutions either broke down or competed for primacy. By integrating the historiographies of Reconstruction, Western history, and Native American ethnohistory this study challenges the notion that federal state formation in the West (and state restoration in the South) were linear processes ushered by a collective of federal actors. Moreover, the existing literature on both Reconstruction and Western territorial expansion has overstated the ability of the federal government to produce communal order through efforts like military occupation, property laws, and multitiered administrative systems such as the Bureau of Indian Affairs. By essentializing the scale of local forces that stacked against federal administration in distant, contested spaces like Montana, the ambitious designs to restore and expand the Union ultimately produced a more exclusionary, unstable, and violent nation.
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Title: Synthesis of HIV-1 GP41 Including FP and MPER by Native Chemical Ligation with Applications to SSNMR. Expression, Solubilization, and Purification of SARS-CoV-2 Spike Protein Subunit 2
Creator: Wolfe, Robert John
Date: 2022
Collection: Electronic Theses & Dissertations
Description: AbstractSynthesis of HIV-1 GP41 Including FP and MPER by Native Chemical Ligation with Applications to SSNMR. Expression, Solubilization, and Purification of SARS-CoV-2 Spike Protein Subunit 2 By Rob Wolfe Human immunodeficiency virus type 1 (HIV-1) and Coronavirus 2019 (Covid-19) have caused substantial risk to public health worldwide. Both HIV-1 and severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) are membrane enveloped viruses which cause acquired immune deficiency...
Show moreAbstractSynthesis of HIV-1 GP41 Including FP and MPER by Native Chemical Ligation with Applications to SSNMR. Expression, Solubilization, and Purification of SARS-CoV-2 Spike Protein Subunit 2 By Rob Wolfe Human immunodeficiency virus type 1 (HIV-1) and Coronavirus 2019 (Covid-19) have caused substantial risk to public health worldwide. Both HIV-1 and severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) are membrane enveloped viruses which cause acquired immune deficiency syndrome (AIDS) and COVID-19 in humans, respectively. The process by which these viruses initiate the infection by membrane fusion of the viral and host cells is similar. However, they differ in which membrane protein is responsible for the fusion process. The HIV-1 glycoprotein 41 (Gp41) is a single-pass integral viral membrane protein containing a ~170-residue ectodomain that is important for membrane fusion between virus and host cells. The ectodomain includes the fusion peptide (FP), N-helical region (NHR), loop, C-helical region (CHR) and viral membrane-proximal external region (MPER). The ectodomain mediates joining (fusion) of the HIV-1 and host cell membranes, which is in an initial step in infection. The ectodomain also adopts different structures, including a final hyper thermostable state. Some data supports a fusion role for this final state. Like HIV-1, SARS2-CoV-2 is enveloped by a membrane that is obtained during budding from an infected host cell. Infection of a new cell requires fusion of the virus membrane with a membrane of the target cell and subsequent deposition of the viral nucleocapsid in the cytoplasm. This process is catalyzed by the Spike (S) protein subunit 2 (S2). My research has focused on the production and characterization of several protein constructs (with or without FP) and the characterization of S2_816-1273, a large S2 protein construct containing all regions including the FP, transmembrane (TM) and the cytoplasmic tail (CT). The expression vector used is pET-24a(+). Biophysical characterization comparisons between S2_816-1273 and shorter constructs including S2_903-998SGGRGG1153-1207 and S2_903-998SGGRGG1163-1207 were performed. Biophysical characterization techniques such as circular dichroism (CD) spectroscopy, size exclusion chromatography (SEC), mass spectrometry (MS), and vesicle fusion assays were used in this study. In addition, this work also describes development and application of native chemical ligation (NCL) methods to produce most of the HIV glycoprotein-41 (gp41) ectodomain. NCL with yields up to 45% were achieved between the chemically synthesized N-terminal FP (gp41512-534) and bacterially synthesized hairpin with MPER (HM) (gp41535-581/SGGRGG/628-683) segments. The hairpin is the NHR-loop-CHR region. These high yields were achieved by solubilizing the HM segment in guanidinium chloride and imidazole which worked together to fully denature the reactants, facilitating the NCL reaction via exposure of the reactant termini. HM was produced with fractional 13C or 2H-labelings that were respectively in the 14-82% and 18% to 54% ranges. The synthesized FP included a non-native N-terminal H6G6D4K tag, which allowed for the binding of FP-HM to a metal-affinity column at the Nterminus to facilitate the separation of unreacted HM. FP-HM was then released from the column by enterokinase-catalyzed cleavage. My work will help future scientists synthesize site-specific 13C and 2H labeled large protein constructs since this is the first time that large Gp41 constructs including full MPER have been synthesized in mg quantities using a combination of bacterial expression system and solid phase peptide synthesis. Furthermore, this methodology is applicable to many proteins that cannot be easily characterized by other methods inside of the lipid bilayer, such as crystallography.
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