Biochemistry, Biophysics, and Structural Biology Commons^™

Host Cell Death In Legionella Pneumophila Pathogenesis And Immunity, Wenhan Zhu

Open Access Dissertations

Legionella pneumophila is an intracellular pathogen that causes a severe, atypical pneumonia termed Legionnaires' disease. Upon entering the host cell, L. pneumophila resides in a membrane-bound vacuole, in which the bacterium evades lysosomal fusion and replicates. The establishment of the vacuole requires the Dot/Icm (Defect in organelle trafficking/ intracellular multiplication) transport system, which translocates a large number of substrates into host cells to re-orchestrate various cellular processes, such as intracellular trafficking, protein synthesis and host cell death pathways. Therefore, a key step in understanding the biology of Legionella is to dissect the mechanisms of action of the Dot/Icm substrates. By …

Associated Behavioral, Genetic, And Gene Expression Variation With Alternative Life History Tactics In Salmonid Fishes, Ashley Chin-Baarstad

Open Access Dissertations

Individual differences in behavior can have potential fitness consequences and often reflect underlying genetic variation. My research focuses on three objectives related to individual level variation: 1) evaluating the innate behavioral variation within and between individuals, families, and progeny of different life-history types across time; 2) testing for differences in gene expression within the brain associated with this behavioral variation; and 3) using genetic polymorphisms to test for associations with ecotype, as well as population structure, in polymorphic populations. First, we evaluated the variation in a suite of ecologically relevant behaviors across time in juvenile progeny produced from crosses within …

Dietary Carbohydrates Influence The Structure And Function Of The Intestinal Alpha-Glucosidases, Mohammad Chegeni

Open Access Dissertations

As the primary products of starch digestion by pancreatic α-amylase, maltooligosaccharides (including maltose) are the main substrates for the α-glucosidases at the intestinal brush border. Here, maltose was shown to induce the formation of a higher molecular weight (HMW) sucrase-isomaltase (SI) species in Caco-2 cells that sorts more quickly to the enterocyte surface to act as a digestive enzyme. As this finding suggested a maltose sensing ability of small intestinal enterocytes, molecular mechanisms associated with the maturation and trafficking of HMW SI were further investigated. A pulse-chase experiment using [ ³⁵S]-methionine revealed a higher rate of early trafficking and …

Towards A Paradigm Shift In The Modeling Of Soil Organic Carbon Decomposition For Earth System Models, Yujie He

Open Access Dissertations

Soils are the largest terrestrial carbon pools and contain approximately 2200 Pg of carbon. Thus, the dynamics of soil carbon plays an important role in the global carbon cycle and climate system. Earth System Models are used to project future interactions between terrestrial ecosystem carbon dynamics and climate. However, these models often predict a wide range of soil carbon responses and their formulations have lagged behind recent soil science advances, omitting key biogeochemical mechanisms. In contrast, recent mechanistically-based biogeochemical models that explicitly account for microbial biomass pools and enzyme kinetics that catalyze soil carbon decomposition produce notably different results and …

Intranuclear Strain Measured By Iterative Warping In Cells Under Mechanical And Osmotic Stress, Jonathan T Henderson

Open Access Dissertations

The nucleus is a membrane bound organelle and regulation center for gene expression in the cell. Mechanical forces transfer to the nucleus directly and indirectly through specific cellular cytoskeletal structures and pathways. There is increasing evidence that the transferred forces to the nucleus orchestrate gene expression activity. Methods to characterize nuclear mechanics typically study isolated cells or cells embedded in 3D gel matrices. Often report only aspect ratio and volume changes, measures that oversimplify the inherent complexity of internal strain patterns. This presents technical challenges to simultaneously observe small scale nuclear mechanics and gene expression levels inside the nuclei of …

Quantum Mechanics In Complex Systems, Ross Douglas Hoehn

Open Access Dissertations

This document should be considered in its separation; there are three distinct topics contained within and three distinct chapters within the body of works. In a similar fashion, this abstract should be considered in three parts. Firstly, we explored the existence of multiply-charged atomic ions by having developed a new set of dimensional scaling equations as well as a series of relativistic augmentations to the standard dimensional scaling procedure and to the self-consistent field calculations. Secondly, we propose a novel method of predicting drug efficacy in hopes to facilitate the discovery of new small molecule therapeutics by modeling the agonist-protein …

The Modification Of Brucine Derivatives As Chiral Ligands And Its Application In The Asymmetric Synthesis, Jian-Yuan Li

Open Access Dissertations

The modification of brucine derivatives as chiral ligands and the use of a multifaceted chiral ligand, brucine diol, under different reaction conditions to produce various optical isomers is described. In Chapter 1, the generation of a number of brucine derivatives is described. Taking the advantage of brucine-diol's excellent molecular recognition capability for multiple organic functional groups, we focused on the synthetic modifications of brucine-diol and the synthesis of brucine N-oxide. We also produced various brucine derivatives with different functional moieties in good yields and selectivities. ^ In Chapter 2, we described the investigation of brucine N-oxide catalyzed Morita-Baylis-Hillman …

New Strategies To Reveal Protein Candidates In Protein-Protein Interactome Study, Meng-Chieh Chen

Open Access Theses

Comprehensive protein-protein interaction network analysis can help reveal protein functions in a system-wide manner. A reliable knowledgebase of interaction networks is not only important for selecting the candidates for drug therapies, but also for evaluating the disease risk. In current interaction databases, 322579 interactions comprised of 56460 proteins have been reported (statistical analysis from APID: Agile Protein Interaction DataAnalyzer; http://bioinfow.dep.usal.es/apid/index.htm). The huge datasets are contributed mainly by yeast -two -hybrid (Y2H) screening and affinity-purification followed by mass spectrometry (AP-MS). High false positive rates and failing to cover certain interaction categories are the limitations of these two methods. Here, we developed …

Transformation Of Biomass Carbohydrates By Transition Metal Catalysts, Christine M Bohn

Open Access Dissertations

By selectively removing functional groups from biomass derived carbohydrates, valuable platform chemicals can be generated from renewable sources. Through dehydration chemistry glucose can be upgraded into 5-(Hydroxymethyl)-2-furfuraldehyde (HMF) and levulinic acid. Iron (III) chloride hexahydrate has shown moderate activity to transform glucose into HMF and has also shown high yields and selectivity for the production of levulinic acid. Typically synthesized from acidic solutions made with mineral acids, levulinic acid has now been produced in high yields with a metal salt. The difference between maximizing production for HMF or levulinic acid from the same catalyst relies on the control of the …

Hierarchical Cell Fluid Extracellular Matrix Interaction In Cell Microenvironment, Soham Ghosh

Open Access Dissertations

Hierarchical structural interactions between components of cell microenvironment, the extracellular matrix (ECM), cytoplasm, nucleus and fluid, are important phenomena that decide cell level physiological process and tissue engineering applications. One of those tissue engineering modalities is freezing of biomaterials, important in a wide variety of biomedical applications including cryopreservation and cryosurgeries. In order to design these applications, freezing-induced changes of the cells and tissues and corresponding biophysical mechanisms need to be well understood. Although the effects of freezing on cells in suspension have been extensively studied, the intracellular mechanics of cells embedded in the extracellular matrix (ECM) during freezing are …

Theory For Diffusional Encounters In Heterogeneous Environments And Multivalent Electrolyte Screening Of Charged Interface, Ran Li

Open Access Dissertations

We develop a theory for encounter rates in a three-dimensional system of connected compartments. The model of connected compartments exhibits the length-scale dependent diffusion that is observed in many heterogeneous environments, such as porous catalysts and biological environments. We discovered a dimensionless number that is the dominant scaling variable and obtained, for the first time, an analytical expression for the encounter rate. The new theory generalizes the classic Smoluchowski diffusion limit to the case of heterogeneous environments. The new theory is tested using Brownian dynamics simulations.^ We also experimentally investigated the behavior of multivalent electrolyte near a charged solid-liquid interface. …

Structure-Functionality Relationship Of Collagen Scaffolds For Tissue Engineering, Seungman Park

Open Access Dissertations

Tissue engineering is a promising technology that enables scientists to create artificial organs or replace damaged tissues using animal cells and other components. For successful tissue regeneration, many factors should be taken into account, however, three components are most crucial: cell, scaffold, and soluble factor(s). In order to check the functionality after regeneration of desired tissues, various approaches have been attempted, depending on the physical, biological, and chemical properties of the tissues. Recently, the importance of the extracellular matrix (ECM) microstructure is being considered to be important in this regard. The ECM is closely associated with various functional properties of …

Understanding Preferred Leg Stiffness And Layered Control Strategies For Locomotion, Zhuohua H. Shen

Open Access Dissertations

Despite advancement in the field of robotics, current legged robots still cannot achieve the kind of locomotion stability animals and humans have. In order to develop legged robots with greater stability, we need to better understand general locomotion dynamics and control principles. Here we demonstrate that a mathematical modeling approach could greatly enable the discovery and understanding of general locomotion principles. ^ It is found that animal leg stiffness when scaled by its weight and leg length falls in a narrow region between 7 and 27. Rarely in biology does such a universal preference exist. It is not known completely …

Effects Of Altered Expression Of The Sumo Conjugating Enzyme, Ubc9 On Mitosis, Meiosis And Conjugation In Tetrahymena Thermophila, Qianyi Yang

Open Access Dissertations

SUMOylation is a critical posttranslational modification in eukaryotic species. Ubc9p is the E2-conjugating enzyme for SUMOylation and consequently it influences multiple cellular pathways. Nuclear proteins are common targets of SUMOylation and regulate nuclear events such as transcription, DNA repair and mitosis. The segregation of the Tetrahymena thermophila genome into two different nuclear compartments provides an unusual context for the analysis of SUMOylation. Each cell contains a transcriptionally silent, diploid germ line micronucleus (MIC) that divides by mitosis and a polyploid transcriptionally active somatic macronucleus (MAC) that divides by an amitotic mechanism. With the long-term goal to exploit these opportunities we …

New Experimental And Theoretical Tools For Studying Protein Systems With Elements Of Structural Disorder, Tairan Yuwen

Open Access Dissertations

Disordered proteins are one class of proteins which do not possess well-folded three-dimensional structures as their native conformations. Many eukaryotic proteins have been found to be fully disordered or contain certain disordered regions. Disordered proteins usually display several characteristic properties, such as increased motional freedom and the conformational heterogeneity caused by that. The elements of structural disorder are commonly involved in many important biological functions and are implicated in many diseases. Therefore, the study of disordered proteins has become one of the most important research topics in recent years. This thesis presents results from three different research projects; the common …

Determining The Binding Between Saga Subunits And Spliceosomal Components, Peyton J. Spreacker, Rachel L. Stegeman, Vikki M. Weake

The Summer Undergraduate Research Fellowship (SURF) Symposium

Proper gene regulation is vital to the health and development of an organism. Determining the relationship between splicing, transcription, and chromatin structure is vital for understanding gene regulation as a whole. There have been previous studies linking these elements pairwise; however, no evidence exists for a direct link between all three. Recent data shows that splicing components of the U2 small nuclear ribonucleic protein (snRNP) co-purify with Spt-Ada-Gcn5-acetyltransferase (SAGA), a highly conserved transcriptional co-activator and chromatin modifier. We hypothesize that SAGA binds with splicing components through a multi-protein binding surface with certain core components based on preliminary yeast two-hybrid data. …

The Effect Of Transient Hmg-Coa Reductase And 1-Deoxy-D-Xylulose-5-Phosphate Synthase Overexpression On Terpene Production In Transgenic Tomato Fruits, Scott A. Gentry, Michael Gutensohn, Laura Henry, Natalia Dudareva

The Summer Undergraduate Research Fellowship (SURF) Symposium

Isoprenoids are secondary metabolites that control numerous plant functions including signaling, growth, photosynthesis, and membrane structure. The bioengineering of isoprenoid synthesis could produce plants with a variety of beneficial traits. Plants form isoprenoids using two different pathways, the mevalonate (MVA) pathway and the methylerithritol phosphate (MEP) pathway, which cooperate via metabolic cross-talk. Transgenic tomato lines expressing both the plastidic and cytosolic forms of the snapdragon nerolidol/linalool terpene synthase under a fruit ripening specific promoter were transiently transformed to overexpress key enzymes in the two isoprenoid pathways. Hydroxymethylglutaryl-coenzyme A reductase (HMGR) is the rate limiting enzyme in the MVA pathway that …

Development Of Electron Microscopy Analysis And Simulation Tools For Nanohub, Mingxuan Lu, Chang Wan Han, Benjamin P. Haley, Volkan Ortalan

The Summer Undergraduate Research Fellowship (SURF) Symposium

Electron microscopy has a crucial role in the field of materials science and structural biology. Although electron microscopy gives lots of important results and findings, some additional simulations and image processing/reconstruction is required to get more information from the data that are collected from the experiments. For this purpose, researchers are using IMOD¹ and QSTEM² for electron microscopy analysis and simulation. IMOD is a set of programs used for tomographic reconstruction and 3D visualization and QSTEM is used for quantitative simulations of TEM and STEM images. However, IMOD and QSTEM are hard to install or use for beginners …

Electrophoresis Staining: A New Method Of Whole Mount Staining, Mitchell G. Ayers, Sarah Calve, Zhiyu Li

The Summer Undergraduate Research Fellowship (SURF) Symposium

Advances in tissue clearing techniques have allowed almost a ten-fold increase in the viewing depth of confocal microscopy. This allows for intact cellular structures to be rendered in 3D. However, viewing tissues to this depth is often limited to endogenous fluorescence as passive diffusion of antibodies via whole mount staining can take weeks. Our lab is developing a new method involving electrophoresis as a driving force that will promote active antibody binding deep into tissue, reducing the amount of time needed to stain for cellular structures. Due to the inherent charge within antibodies, they are able to be directionally forced …

Abcb11 Functions With B1 And B19 To Regulate Rootward Auxin Transport, Jesica Elyse Reemmer

Open Access Theses

Auxin transport is essential for the architecture and development of erect plants. In a network of transporters directing auxin flows, ATP-Binding Cassette (ABC) transporters are a ubiquitous family of proteins that actively transport important substrates, including auxins, across the plasma membrane. ABCB1 and ABCB19 have been shown to account for the majority of rootward auxin transport, but residual fluxes to the root tip in Arabidopsis b1b19 double mutants implies the involvement of at least one additional auxin transporter in this process. Of specific interest, the severe dwarfism seen in abcb1abcb19 is strikingly reminiscent of that seen in mutants defective in …

Efficacy Of Cleaning Method For Removal Of Exogenous Welding Fume Contamination From Nail Tissue Prior To Use As A Biomarker For Welding Fume Manganese Exposure, Jeffrey Corral Bainter

Open Access Theses

Nail tissue has been proposed as a biomarker for body burden of occupational exposure to manganese from welding fumes. Though recent studies have shown correlation between manganese exposure and both nail tissue concentration as well as concentrations in dopaminergic regions of the brain, concerns of the validity of nail tissue as a biomarker have arisen due to the potential for exogenous contamination of Mn to undermine the quantization of endogenous Mn in nail. Previous studies have used a cleaning method of 1% Triton X-100 surfactant plus sonication in order to attempt to remove exogenous welding fume contamination. Determination of the …

1,25-Dihydroxyvitamin D Alters Lipid Metabolism And Epithelialto- Mesenchymal Transition In Metastatic Epithelial Breast Cancer Cells, Alle Nicole Barnard

Open Access Theses

Evidence suggests that high vitamin D status (marked by serum 25-hydroxyvitamin D, 25(OH)2 D) is associated with a decreased risk of breast cancer. It has been established that 1,25-dihydroxyvitamin D (1,25(OH) 2 D) can alter glycolysis and the Krebs cycle of breast cancer cells (Jiang et al., 2010; Zheng et al., 2013) but little information is available on 1,25(OH) 2 D's alterations of lipid metabolism in breast cancer cells. Thus, the current research investigates if there was an effect of 1,25(OH) 2 D on proteins that regulate lipid metabolism in MCF10A, MCF10A-ras, MCF10CA1h, and MCF10CA1a epithelial breast cancer cells. While …

Using The Microsoft Kinect To Assess Human Bimanual Coordination, Joshua James Liddy

Open Access Theses

Optical marker-based systems are the gold-standard for capturing three-dimensional (3D) human kinematics. However, these systems have various drawbacks including time consuming marker placement, soft tissue movement artifact, and are prohibitively expensive and non-portable. The Microsoft Kinect is an inexpensive, portable, depth camera that can be used to capture 3D human movement kinematics. Numerous investigations have assessed the Kinect's ability to capture postural control and gait, but to date, no study has evaluated it's capabilities for measuring spatiotemporal coordination. In order to investigate human coordination and coordination stability with the Kinect, a well-studied bimanual coordination paradigm (Kelso, 1984, Kelso; Scholz, & …

Investigation Into The Control Of Melittin Secondary Structure And Antimicrobial Activity, Zachary B. Molinets

Open Access Theses

Antimicrobial resistance has been an exponentially growing problem since the discovery of antibiotics. Antibiotics have been misused for many years and this misuse has grown into a real problem for the medical community. While there are countless safeguards to prevent infection by a resistant strain of bacteria, there are still many plagued by it and must be treated with sometimes dangerous antibiotics. Melittin, along with many other peptides, contain potent antimicrobial properties, but are also toxic toward enthrocytes. The control of the secondary structure of peptides provides the key to adjusting their activity.

Key Residues Of Human Cytoplasmic Protein Tyrosine Phosphatase-A And -B For Substrate Binding And Specificity, Byunghyun Park

Open Access Theses

Reversible tyrosine phosphorylation plays an important role in signaling pathways that are essential for regulating cellular growth, differentiation and metabolism. Moreover, several human diseases such as diabetes, obesity and cancers are associated with the deregulation of protein tyrosine phosphatases (PTPs). Several studies provide evidence that PTPs not only contribute to cellular differentiation, but over-expression of these molecules also leads to transformation of non-transfomed cells as well. Based on these results, designing specific PTP inhibitors may ultimately function as potential therapeutic agents to treat various diseases including cancer, diabetes, and autoimmune diseases. EphA2 is a receptor tyrosine kinase which is hypo-phosphorylated …

Synthesis And Dna-Binding Studies With Two Stericallyfriendly Porphyrin Frameworks, Srijana Ghimire

Open Access Dissertations

Peripheral substituents on cationic porphyrins play a significant role during binding with DNA hosts. Possible applications of these systems in photodynamic therapy as well as in anti-bacterial and anti-cancer therapies motivate the binding studies. For characterizing DNA binding motifs different methods are useful including absorption, emission, and circular dichroism spectroscopies, as well as viscometry and X-ray crystallography. With the classic H₂ T₄ porphyrin, or 5,10,15,20-tetra(N-methylpyridinium-4-yl)porphyrin, the mode of binding varies with the base composition of the DNA host. The porphyrin binds adenine-thymine rich sequences externally whereas intercalation occurs in guanine-cytosine rich sequences. The McMillin group has made some …

Metal Stopping Reagents Facilitate Discontinuous Activity Assays Of The De Novo Purine Biosynthesis Enzyme Pure, Kelly L. Sullivan, Loredana C. Huma, Elwood Mullins, Michael E. Johnson, T. Joseph Kappock

Department of Biochemistry Faculty Publications

The conversion of 5-aminoimidazole ribonucleotide (AIR) to 4-carboxy-AIR (CAIR) represents an unusual divergence in purine biosynthesis: microbes and nonmetazoan eukaryotes use class I PurEs while animals use class II PurEs. Class I PurEs are therefore a potential antimicrobial target; however, no enzyme activity assay is suitable for high throughput screening (HTS). Here we report a simple chemical quench that fixes the PurE substrate/product ratio for 24 h, as assessed by the Bratton-Marshall assay (BMA) for diazotizable amines. The ZnSO₄ stopping reagent is proposed to chelate CAIR, enabling delayed analysis of this acid-labile product by BMA or other HTS methods

Developing An Unstructured Model To Investigate The Effect Of Ethanol On Product Yields For Glucose And Xylose Cofermentation In Saccharomyces Cerevisiae 424a (Lnh-St), Shane D. Clingenpeel

Open Access Theses

Production of bio-ethanol from lignocellulose requires the efficient fermentation of glucose and xylose, even in the presence of inhibitors. The desired product, ethanol itself, will inhibit the fermentation. A further understanding of how ethanol affects the organism is critical to overcoming its inhibition.

This thesis evaluated the effect of ethanol on the cofermentation of glucose and xylose in two different cases. The first case had an unstructured model created for Saccharomyces cerevisiae 424A (LNH-ST), a genetically modified strain of yeast capable of cofermenting glucose and xylose. The differential equations were based around sugar consumption, and the product yields were investigated …

Neurotrophins And Their Effects On Breast Cancer Cell Proliferation And Migration, Kayla Elise Minser

Open Access Theses

Cancer is a large health issue in all parts of the world. In the United States alone, approximately 1 in 4 deaths are cancer related. Breast cancer is a particularly prevalent form, accounting for a little over 14 percent of all cancer incidence. The largest obstacle to overcome for breast cancer morbidity is metastasis. Over 90 percent of all breast cancer related deaths are due to metastasis. Because metastasis is a complex, multi-step process, it is difficult to treat. A recent observation in the Kirshner lab has revealed a type of phenotypic plasticity, where migratory cancer cells have a neuronal-like …

Pharmaceuticals And Personal Care Products: Emerging Contaminants In Aquatic Ecosystems, Jenny E. Zenobio

Open Access Theses

In recent years, the presence of pharmaceuticals and personal care products (PPCPs) in aquatic systems has led to research on their fate and effects. PPCPs have been found in mixture in wastewater effluents, surface, ground, and drinking water at low concentrations from areas of intense urbanization. Although adverse effects to human health from the current environmental concentrations are unlikely, the impacts to ecological receptors are not clear. We performed field and laboratory studies to quantify and evaluate effects of PPCPs on fish. First, a field study was conducted at the Baca National Wildlife Refuge, Colorado (2010-2012) because a portion of …