Biochemistry, Biophysics, and Structural Biology Commons^™

Vertical Transmission Of Zika Virus In An Opossum Model, Dionn Carlo Silva

Theses and Dissertations

This study will identify Brazilian ZIKV infection in the offsprings of mated, inoculated Monodelphis domestica females. Unlike non-human primates (NHP) and transgenic mice, M. domestica provides an opportunity to study ZIKV infection in a novel animal model possibly useful for in-utero studies. M. domestica has also been used in cancer and dietary research with varying degrees of success. ZIKV infection will be identified using antibody staining and in situ hybridization. The expected result of this study is that ZIKV will be identified in various tissues and reveal that ZIKV is transmitted vertically in this animal model. The significance of this …

Developmental Activation Of Mitochodnrial Opa1 Processing Is Retinoic Acid-Independent, Shaynah St. Vallier

Theses and Dissertations

Mitochondria exist as an organellar network within most eukaryotic organisms, and are critical to biochemical energy production and cellular stress response. Mitochondrial bioenergetic function is directly linked to the complex morphological state of its network, existing in a balance of fusion (interconnected state) or fission (fragmented state). The optic atrophy-1 (OPA1) protein plays a major role in regulating mitochondrial inner membrane fusion, whereas the OMA1 metallopeptidase is highly involved in the degradation of the OPA1 protein, thus contributing to its fragmented state. Cellular stress such as a disruption in mitochondrial membrane potential activates OMA1 cleavage of OPA1. However, our data …

A Computational Investigation Of The Biophysical Mechanisms Underlying Thermotaxis In The Afd Neurons Of Caenorhabditis Elegans, Zachary Mobille

Theses and Dissertations

Thermotaxis in the nematode Caenorhabditis elegans (C. elegans) is studied at the cellular scale of the amphid finger-like ciliated (AFD) neurons, which have previously been shown to be essential for thermoreception. The voltage and calcium signals of AFD during temperature stimuli are described with ordinary differential equations. The primary calcium model is a modified version of that published by Kuramochi and Doi in 2017 to explain the calcium responses of the chemosensitive amphid single-ciliated right (ASER) neuron to fluctuations in extracellular salt concentration. To account for the effects of temperature, changes to the stimuli conditions under which inactivation takes place …

Reducing Seed Coat Fiber Content And Pod Shatter, And Engineering Medium Chain Fatty Acids-Containing Oil, In The Oilseed Crop Pennycress ( Thlaspi Arvense L. ), Maliheh Esfahanian

Theses and Dissertations

The overall goal of this thesis was to genetically improve agronomic traits of pennycress (Thlaspi arvense L.; Field Pennycress) and demonstrate the production of value-added designer seed oils to domesticate pennycress and enable its establishment as a new winter annual oilseed/meal/cover crop to be grown in temperate regions of the world. In the U.S. Midwest, pennycress can be double cropped on existing farmland during the time between corn harvest and subsequent planting of soybeans the following spring. Pennycress has the potential to produce 2,000 lbs/acre seeds, which at 33% by weight oil content and 20% protein, would yield 85 gallons/acre …

Design Of Fibril Forming Collagen Mimetic Peptides: Heterotrimers And Nucleation Domains, Sally Tan

Theses and Dissertations

This paper attempts to design collagen mimetic peptides where the triple-helical region mimics that of human Type I Collagen. With consideration for chain selection and chain register, we utilize the NC2 domain of heterotrimeric Type IX Collagen as a nucleation domain for triple-helix folding.

Influence Of Metal Sleeves In The Accuracy Of Dental Implant Placement Using Guided Implant Surgery, Coleman Adams

Theses and Dissertations

The fabrication of implant surgical guides through stereolithographic 3D printing has become a staple in dental implant guided surgery over the last couple decades. These surgical guides have typically utilized metal sleeves to assist in guidance of the drills during osteotome preparation. The metal sleeves can be costly and potentially cause deviations if improperly placed during post-processing of the guide. This research explored a novel method for the utilization of sleeve-free surgical guides by comparing the dimensional and angulational deviations between the implant guides with and without a metal sleeve. To achieve this goal, two separate aims were pursued. Our …

Computational Analysis And Prediction Of Intrinsic Disorder And Intrinsic Disorder Functions In Proteins, Akila I. Katuwawala

Theses and Dissertations

COMPUTATIONAL ANALYSIS AND PREDICTION OF INTRINSIC DISORDER AND INTRINSIC DISORDER FUNCTIONS IN PROTEINS

By Akila Imesha Katuwawala

A dissertation submitted in partial fulfillment of the requirements for the degree of Engineering, Doctor of Philosophy with a concentration in Computer Science at Virginia Commonwealth University.

Virginia Commonwealth University, 2021

Director: Lukasz Kurgan, Professor, Department of Computer Science

Proteins, as a fundamental class of biomolecules, have been studied from various perspectives over the past two centuries. The traditional notion is that proteins require fixed and stable three-dimensional structures to carry out biological functions. However, there is mounting evidence regarding a “special” class …

A Novel Review Of Heat Shock Protein 110 Kda: A Basis For Research And Continued Experimentation Through Biochemical Analysis, Crist W. Cuffee

Theses and Dissertations

Heat shock protein 110 kDa, Hsp110, is a distinct cellular protector, different in form and function from Hsp70, a close relative of Hsp110. Functioning primarily as a holdase or in tandem with other molecular chaperones, a review of current accomplishments elucidates the uniqueness of this protein and the continued mysteries that surrounds it. Found only in eukaryotes, Hsp110 has been linked to many diseases, ranging from parasitic infection to neurodegenerative disorders. While still lacking, studies of this protein have provided much in the realm of speculation on the mechanisms behind Hsp110s’ contribution to different pathologies. This review will serve as …

Single Molecule Investigations Of Holliday Junction Binding Protein Ruva, Dalton Reed Gibbs

Theses and Dissertations

DNA breaks are inevitable as they mainly occur due to cells’ own reactive oxygen species (ROS). While DNA breaks can be single-stranded or double-stranded, the double-stranded DNA (dsDNA) breaks are more dangerous. If such damage is not repaired, it can lead to genetic instability and serious health issues including cancers. One way dsDNA breaks can be repaired is via a process called homologous recombination (HR), which involves several DNA-binding proteins. Therefore, to have a better insight into the repair mechanism and origin of repair defects, we need a better understanding of how these proteins interact with DNA itself and DNA …

Novel Roles Of Yy1 In Astrocytes, Karli A. Mockenhaupt

Theses and Dissertations

Astrocytes tile the entire central nervous system, are diverse across brain regions, and perform specialized functions fine-tuned for their neuronal circuit. Initially, astrocyte heterogeneity is generated through intrinsic programs specified by their progenitors. Then, as astrocyte mature, they are specialized by region-specific communication with surrounding cells. The molecular programs that regulate this diversity and allow for the accommodation to the needs of the local environment remains elusive. We explored a ubiquitously expressed zinc finger transcription factor, Yin Yang 1 (YY1), that both activates and represses transcription by regulating loops of active chromatin. This report describes the diverse functions of astrocytic …

Muscarinic Excitation Of Dopamine Neurons In The Ventral Tegmental Area Via Activation Of A Trpc-Like Cation Conductance, Yu Tzu Chen

Theses and Dissertations

Dopaminergic (DA) neurons in the ventral tegmental area (VTA) play a crucial role in reward and motivational behaviors, including the development of drug addictions. VTA DA neurons receive excitatory cholinergic inputs from the mesopontine tegmentum. Blockage of the M5 muscarinic receptor in DA neurons has been shown to attenuate drug-induced DA release and abuse-related behaviors, but the molecular mechanism is unknown. In this study, experiments were designed to identify the electrophysiological effects of muscarinic agonism in the modulation of action potential kinetics and firing patterns in VTA DA neurons of mice. Pharmacology of the muscarinic receptor-evoked current was also characterized. …

Lonidamine Is A Novel Inhibitor Of Fatty Acid Oxidation, Tianhai He

Theses and Dissertations

In this thesis project, we examined the hypothesis that lonidamine is an effective inhibitor of FAO in mammalian cells, underling its inhibitory effects on bioenergetic production and anti-cancer action. We compared its effects on FAO and glycolysis in multiple human cancer cell lines of different lineages.

Glycosaminoglycan Modulation Of Key Proteins In Alzheimer's Disease, Tamim Chiba

Theses and Dissertations

Glycosaminoglycans (GAGs) are linear polysaccharides whose disaccharide building blocks consist of an amino sugar (D-glucosamine) and either uronic acid (D-glucuronic acid or L-iduronic acid) or galactose. Nearly all GAGs (with the exception of hyaluronic acid) are covalently attached to proteins, forming proteoglycans, and are ubiquitous in nature. The monomeric units, chain length, degree and pattern of sulfation, and epimerization are important properties that determine the influence of these molecules. Through Coulombic forces, hydrogen bonds and hydrophobic interactions, they regulate many physiological and pathophysiological processes such as blood coagulation, cell growth and differentiation, inflammatory processes, host defense and viral infection mechanisms. …

Mechanistic Insights Of Mitochondrial Reactive Oxygen Species In Alzheimer's Disease Models, Jakob C. Green

Theses and Dissertations

Alzheimer’s disease (AD) is a neurodegenerative disease that results in impaired cognition, disorientation, confusion, poor judgement, and behavioral changes. The mitochondrial cascade hypothesis (MCH), which postulates that accumulating oxidative stress and mitochondrial dysfunction plays an integral role in the development of the disease, potentially upstream, in conjunction with or independent of Aβ and tau. The literature supports the critical role of mitochondrial dysfunction in the progression and development of AD, but is inconclusive about what precise role, as initiator or delegate, it performs. Regardless, the impact that oxidative stress and mitochondrial dysfunction have on AD deserves further study and investigation. …

Cryogenic Electron-Microscopy (Cryo-Em) Analysis Of Mscs, Ynai, Connexin 26 And Bioinformatic Analysis Of Cysteine And Serine In Experimental Protein Structures, Claudio Catalano

Theses and Dissertations

Functional insights into macromolecules lie in the arrangement of their molecular structures. Investigating the atomic framework of a biomolecule could unwind its biochemical mechanism, providing aid to many relevant biological questions, thus facilitating drug discovery processes. Two approaches have been used to gain knowledge at the atomic level of a protein function: experimental and computational methodologies.

From an experimental perspective, membrane proteins (MPs) were investigated. Such proteins represent ~20% of the human proteome, and ~60% of them are drug targets since they are exposed to the surface of cells and are responsible for a wide range of fundamental functions like …

Characterization Of Cytochrome P450 3a4 Structure And Ligand Binding Mechanism In Anionic Nanodiscs, David T. Sweeney

Theses and Dissertations

The Cytochromes P450 superfamily are ubiquitous throughout life and are present in some viruses. Humans encode for 57 of these enzymes that are involved in the metabolism of ~90% of all chemicals. These membrane-bound monooxygenase enzymes utilize a heme protoporphyrin at their active site to activate molecular oxygen and oxidize their substrates through an intermediate deemed nature's most powerful oxidizer. They are a powerful system at our disposal yet understanding of their structure and ligand binding mechanisms in the presence of a lipid bilayer remains poorly understood. The focus of this work is on Cytochrome P450 3A4 (CYP3A4). We set …

Development Of Unnatural Amino Acid Mutagenesis Technologies To Control Enzyme Activity And Synthesis Of Chemically Reactive Ubiquitin Probes, Jenny Cheung

Theses and Dissertations

Most of the work vital for the survival of the cell are performed by proteins. The primary sequence of proteins is composed of 20 canonical amino acids. However, these 20 amino acids do not offer the full range of functionalities needed for proteins to do their job. Often times, post-translational modifications (PTM’s) occur to add the required functional group. This interaction between all the different proteins in the proteome in addition to PTM’s create a complex system that scientists are still trying to unravel to this day. Many different tools had been developed for this purpose, and one such tool …

Interclass Gpcr Heteromerization Affects Localization And Trafficking, Rudy Toneatti

Theses and Dissertations

Class A serotonin (5-hydroxytryptamine) 2A (5-HT_2AR) and class C metabotropic glutamate 2 receptors (mGluR2) are seven transmembrane receptors (7TMRs or G protein-coupled receptors – GPCRs) involved in multiple neuropsychiatric disorders including schizophrenia. Previous findings from our laboratory reported that 5-HT_2AR and mGluR2 are dysregulated in the prefrontal cortex of patients suffering from this psychiatric condition, although 5-HT_2AR’s expression was recovered in antipsychotic-medicated patients. Genome-wide association studies on schizophrenia reported that endosomal trafficking that regulates cell surface abundance of another 7TMR implicated in this disease (dopamine D2 receptor) can be altered. Ligand-activated receptors, including the …

Development And Evaluation Of A Combinatorial Rt-Qpcr Multiplex For Forensic Body Fluid Identification, Carolyn A. Lewis

Theses and Dissertations

Body fluid identification is essential in the forensic biology workflow that assists DNA analysts in determining where to collect DNA evidence. Current presumptive tests lack the sensitivity and specificity molecular techniques can achieve; therefore, molecular methods, such as microRNA and microbial signatures, have been extensively researched in the forensic community. Limitations of each method suggest combining molecular markers to increase discrimination efficiency of multiple body fluids from a single assay. While microbial signatures have been successful in identifying fluids with high bacterial abundances, microRNAs have shown promise in fluids with low microbial abundance. A disadvantage of RNA analysis in forensic …

Detrimental Role Of Irf1 In Myeloid Cells During Influenza Infection, Bianca L. Baum

Theses and Dissertations

The influenza virus is constantly evolving new ways to overcome both innate and vaccine-induced host defenses, making the innate immune response a critical therapeutic target for minimizing morbidity and mortality caused by infection. Here we examined the role that interferon regulatory factor 1 (IRF1) plays in regulating host innate immune responses in myeloid cells. We found that knocking out IRF1 from myeloid cells in mice (IRF1^DMy) led to a partial protection during IAV infection. Interestingly, recruitment of bone marrow derived myeloid cells as well as NLRP3 inflammasome expression were decreased in the lungs of IRF1^DMy mice, whereas …

Virtual Reality Tool For Visualizing The Oral Microbiome, Alexus M. Harris

Theses and Dissertations

There are limited teaching tools for helping both students and patients understand the oral microbiome. It is important to understand the oral microbiome as it heavily influences the health of your oral cavity and the dysbiosis of disease. The oral microbiome influences other aspects of health and serves as an indicator for health. With a greater understanding of the oral microbiome, patients can make healthier lifestyle changes and dentists can better educate their patients. For that reason, the goal of this project was to create a virtual reality tool for visualizing the oral microbiome. The assets for the VR tool …

Conformational Modulation Of Α-Helical N-Acetylated Α-Synuclein, Ricardo Fernandez

Theses and Dissertations

Human N-acetylated α-synuclein (^NAcαSyn) has long been accepted as an intrinsically disordered protein (IDP) predominantly found in terminal nerve regions in healthy subjects. ^NAcαSyn structural fluidity enables the formation of transient or dynamic assemblies. Some of these assemblies may serve functional, yet undefined purposes. ^NAcαSyn is highly susceptible to the surrounding environment, leading to misfolding under high cytosolic oxidative stress conditions, which in turn assemble in the form of toxic oligomers or amyloid fibrils. These ^NAcαSyn inclusions along with their accretion and migration within neurons, are pathological hallmarks of Parkinson’s disease (PD), dementia, and multiple …

A Mechanism Behind The Mechanotransduction Of Surface Characteristics In Osteoblasts, Otto J. Juhl Iv

Theses and Dissertations

Biomaterials for use in bone regeneration and healing range from metal and metal alloy implants to hydrogel-based solutions. These materials can be optimized to increase bone healing and integration by improving the mechanical and biological properties. Regardless of the material itself, the cell-substrate interaction is key to the success of the biomaterial once implanted. Substrate surface characteristics such as roughness, wettability, and particle density are well-known contributors to a substrate’s overall osteogenic potential, and therefore the substrate's overall success. Unfortunately, it is still unknown how these substrate surface characteristics are transduced into intracellular signals by cells, preventing specific tailoring of …

A Molecular Toolset For The In Vivo Detection Of A Sulfolobus Islandicus Leucyl Trna Synthetase Paralog, Nicholas Michael Bretz

Theses and Dissertations

An ancient and ubiquitous set of enzymes known as the aminoacyl-tRNA synthetases are required for the viability of all organisms. The aminoacyl-tRNA synthetases catalyze the attachment of amino acids onto tRNA molecules. The aminoacylated, or charged, tRNA is then transported and utilized at the ribosome for the synthesis of proteins. The genome of the hyperthermophilic microorganism Sulfolobus islandicus (S. islandicus) harbors a unique leucyl-tRNA synthetase paralog, LeuRS-I, of unknown function. While its duplicate, LeuRS-F, carries out all the classical functions of LeuRS-family enzymes, LeuRS-I fails to charge leucine onto tRNALeu, despite its ability to activate this amino acid and bind …

Stem Cell Technology For Age Related Macular Degeneration Intervention, Laura Valdez

Theses and Dissertations

The main focus of this thesis is to study RPE cell derived from IMR90 induced pluripotent stem cells in various physiological conditions mimicking RPE transplantation rejection. Retinal pigmented epithelium (RPE) cells are located between the choroid and photoreceptors in the eye and they are essential to provide nutrients from blood to rods and cones, as well retinoids of the visual cycle. Vision loss and various ocular diseases are attributable to the degeneration or dysfunction of the RPE cells, leading to blindness. One of the major ocular problem from RPE dysfunction is macular degeneration. Age-related macular degeneration (AMD) can be frequently …

Snow-Albedo Feedback In Northern Alaska: How Vegetation Influences Snowmelt, Lucas C. Reckhaus

Theses and Dissertations

This paper investigates how the snow-albedo feedback mechanism of the arctic is changing in response to rising climate temperatures. Specifically, the interplay of vegetation and snowmelt, and how these two variables can be correlated. This has the potential to refine climate modelling of the spring transition season. Research was conducted at the ecoregion scale in northern Alaska from 2000 to 2020. Each ecoregion is defined by distinct topographic and ecological conditions, allowing for meaningful contrast between the patterns of spring albedo transition across surface conditions and vegetation types. The five most northerly ecoregions of Alaska are chosen as they encompass …

Analysis Of Olfactory Mrna Cap Modifications Using Rnaseh And Lc-Ms, Ilinca M. Giosan

Theses and Dissertations

In the olfactory system, olfactory sensory neurons (OSNs) express odorant receptors (ORs) in a singular fashion through unknown mechanisms. Published data supports the notion that singularity is achieved, in part, through a modification the mRNA strand, perhaps in its 5’cap region. This theoretical study aims to test this 5’cap hypothesis.

Adsorption Behavior Of Chemically / Charged Modified Antibodies On Gold Nanoparticles, Samuel Okyem

Theses and Dissertations

Gold nanoparticles (AuNPs) have been exploited in the various domains of science such as drug delivery, bio-sensing, immunoassays and environmental sensors, due to their optical properties and intriguing surface chemistry. Different scientific procedures have been used to effectively immobilize antibodies onto AuNPs. Although acceptable outcomes have been achieved in the immobilization of antibodies onto AuNPs, the sensitivity of these immobilized antibodies to target antigen or binding sites is limited due to improper orientation of the antibodies. Also, the possibility of nanoparticle aggregation when exposed to proteins limits its biomedical applicability.

There is some evidence that the surface charge of antibodies …

Examining Various Aspects Of Zika Virus Dissemination In Aedes Aegypti, Ileana C. Lozano

Theses and Dissertations

South Texas is one of the few locations where Zika virus has been locally transmitted in the U.S. It has a climate which is distinct to other areas with autochthonous transmission, with extremely hot, dry summers and moderate winter temperatures. Studying mosquito transmission of Zika in a range of temperatures conditions replicating where virus transmission is occurring is essential in order to have a better understanding of transmission patterns. These factors were examined by infecting mosquitoes and monitoring the dissemination status through real-time PCR analysis.

To further characterize dissemination of Zika virus within south Texas mosquitoes a time series immunohistochemical …

Acetoacetate Enhances Oxidative Metabolism And Response To Toxicants Of Cultured Renal Cells, Trudi Maria Denoon

Theses and Dissertations

In vitro methods can be cost effective and facilitate large scale drug screening in the pharmaceutical industry. Renal proximal tubular cells have been a prime target for drug induced toxicity. LLC-PK1 cells possess many transport functions of the proximal tubule epithelia, however, its energy metabolism differs from the highly oxidative tubule metabolism in vivo. This may impact its reliability in predicting toxicity. Modulating the growth media composition has been shown to shift cultured liver and muscle cells from high glycolytic activity to increased oxidative metabolism, but few studies have been done on renal cells. This study is designed to test …