Biochemistry, Biophysics, and Structural Biology Commons^™

Pre-Administration Of Medium Chain Triglycerides In Vivo Can Attenuate Or Block The Effects Of Recurrent Hypoglycemia, Dhanisha Nandigama

Biological Sciences

Hypoglycemia is a state of abnormally low blood glucose. Many patients who use insulin, primarily for the treatment of diabetes, experience multiple bouts of hypoglycemia, termed recurrent hypoglycemia (RH). Because RH impairs cognitive function and ability to appropriately respond to a subsequent episode of hypoglycemia, it is critical to develop treatments. One approach, which we have taken here, is to attempt to preserve neuronal fuel supply during a hypoglycemic episode. Medium-chain triglycerides are medium-chain fatty acid (MCT) esters of glycerol that can provide an alternative fuel source to the brain via ketones; the hippocampus is known to express transporters for …

Relationships Of Protein Biomarkers Of The Urokinase Plasminogen Activator System With Expression Of Their Cognate Genes In Primary Breast Carcinomas., Seth B. Sereff

College of Arts & Sciences Senior Theses

Background: Urokinase plasminogen activator (uPA), its receptor uPAR and serine protease inhibitors PAI-1 or PAI-2 play key roles in tissue membrane remodeling and invasion of basement membranes by induction of a fibrinolytic pathway. Earlier studies reported that uPA and PAI-1 protein levels in breast carcinomas assist in prediction of response to chemotherapy. Our goal is to develop molecular signatures of candidate genes and identify novel relationships with these four protein biomarkers that demonstrate clinical utility for assessment of breast carcinoma outcomes.

Methods: This retrospective study used de-identified biomarker results and clinical outcomes from primary breast cancers that were stored in …

Dom Control Of Mercury Methylation In The Water Column Of A Meromictic Lake, Madalyn Bozinski

Renée Crown University Honors Thesis Projects - All

The trophic transfer and bioaccumulation of methyl mercury (MeHg) in aquatic ecosystems is a substantial concern, resulting in fish consumption advisories worldwide. Aquatic ecosystems have been identified as the critical environments that breed production of MeHg and low levels of initial accumulation in biota. MeHg production is a microbially-mediated process, occurring primarily at the transition between oxic and anoxic environments. This research aimed to assess the extent to which dissolved organic matter (DOM) affects methylation of mercury in the oligotrophic meromictic lake, Green Lake, in Fayetteville, NY and the warm monomictic lake, Seneca Lake, in Geneva, NY. General physical parameters …

An Exploration Of Ruthenium Catalyzed Alder-Ene Reactions, Emily Bonner

Renée Crown University Honors Thesis Projects - All

This project focuses on expanding the reaction scope of the ruthenium (Ru) catalyzed Alder-ene reaction with both exocyclic enol ethers and simple alkenes. Knowledge gained from these reactions may be applied to the development of efficient methods for the synthesis of both oxygen heterocycles and acyclic compounds that incorporate a 1,4-diene unit. Each of the two double bonds in the diene possesses different reactivity which increases the utility of these products as synthetic intermediates by allowing for greater versatility in synthesizing natural product targets. A breadth of alkyne and alkene couplings were attempted in this project via the Ru catalyzed …

Reactive Glutamines And A Binding Site Region Contribute To Fxiii Substrate Specificity For Fibrinogen Αc, Mohammed Hindi

College of Arts & Sciences Senior Theses

The reaction between transglutaminase factor XIII (FXIII) and fibrin is a key process in the final stages of blood coagulation. The αC region of fibrin contains three reactive glutamines (Q237, Q328, and Q366) which are crosslinked by FXIII, as well as a FXIII anchoring site (E396). Previous work has helped elucidate the FXIII-αC interaction, but much is still unknown about the key residues in the αC region.

Four crosslinking assays were carried out to characterize the contribution of the αC anchoring site and reactive glutamines in crosslinking by FXIII. The effects of individual E396A, Q366N, and Q328P (Fibrinogen Seoul II …

Autofluorescence To Study The Effects Of Acid Concentration On Cellular Metabolism In Vitro, Robin L. Raley

Chemistry & Biochemistry Undergraduate Honors Theses

Ultraviolet (UV) radiation-induced sunburns and their accompanying afflictions are a growing public health concern in the United States. There is a need for techniques that can accurately and non-invasively characterize the physiology of sunburned skin tissue directly after UV-damage and applying a topical skin treatment to relieve pain and promote healing. Two-photon excited fluorescence (TPEF) microscopy and fluorescence lifetime imaging (FLIM) can be used to investigate metabolic processes in live cells through endogenous fluorescence of the cofactors, NADH and FAD. These methods employ the optical redox ratio of FAD/(NADH+FAD), mean NADH lifetime, and the separation of the free and bound …

Protein Engineering For Biomedical Applications, Claudia Heritage

Renée Crown University Honors Thesis Projects - All

The Korendovych lab recently demonstrated for the first time that short peptides can selfassemble in the presence of metal ions to produce efficient catalysts in a variety of chemical reactions ranging from ester hydrolysis to oxygen activation. Original catalysts were produced by solid phase peptide synthesis. Solid phase synthesis can become the limiting factor for an experiment because it takes over two days to make and purify one peptide. Depending on characteristics of a peptide, the purification process can become a challenge involving a tedious process of protocol optimization. Screening of several peptides is time-consuming as it involves analysis of …

The Synthesis, Purification, And Characterization Of The P3 Peptoid, Myles Joyce

Chemical Engineering Undergraduate Honors Theses

The relatively young field of nano-systems has vast applications over various different fields. Peptoids (peptide analogues) and their uses are consistently investigated components of these nano-studies. A particularly interesting group of peptoids are able to self-assemble into secondary structures under the right conditions. The P3 peptoid is able to form microspheres if properly synthesized and purified. Microspheres are frequently studied due to their low surface area to volume ratios. This unique aspect of microspheres makes them an excellent candidate for drug delivery systems; however, these systems must also be robust so that the desired compound can be encapsulated within the …

The Chelation Of Metal Ions By Vicibactin, A Siderophore Produced By Rhizobium Leguminosarum Atcc 14479, Joshua Stinnett

Undergraduate Honors Theses

Vicibactin is a small, high-affinity iron chelator produced by Rhizobium leguminosarum ATCC 14479. Previous work has shown that vicibactin is produced and secreted from the cell to sequester ferric iron from the environment during iron-deplete conditions. This ferric iron is then transported into the cell to be converted into ferrous iron. This study uses UV-Vis spectroscopy as well as ion trap-time of flight mass spectroscopy to determine that vicibactin does form a complex with copper(II) ions, however, at a much lower affinity than for iron(III). Stability tests have shown that the copper(II)-vicibactin complex is stable over time. The results of …

Transformation Of Model Organisms With Munc13-1 For Subcloning And Protein Harvesting, Grace Dittmar

Honors Theses

Exocytosis is the process by which cells release various molecules into the extracellular space. This mechanism is accomplished by membrane fusion, in which the membrane of the transport vesicle and that of the cell itself fuse to create one continuous membrane, allowing the release of the molecules inside the vesicle. A family of proteins, known as SNARE proteins, is responsible for facilitating membrane fusion. SNARE proteins form a complex between the vesicle membrane and the plasma membrane. For various cells, such as mast cells and neurons, exocytosis is essential to carry out their vital functions. Various accessory proteins are known …

The Dna Damage Response Gene Ddr48 Regulates Sterol Synthesis Genes To Confer Antifungal Resistance To The Pathogenic Fungus Histoplasma Capsulatum, Gabriella Reyes

Honors Theses

Histoplasma capsulatum is a dimorphic fungal pathogen that is endemic to the Ohio, Missouri, and Mississippi river valley regions. The fungus grows as a mold at environmental temperatures (25C) and transforms into a unicellular yeast upon inhalation by a mammalian host (37C). The mold to yeast shift is required for pathogenicity in host organisms, where the potentially fatal disease, histoplasmosis, can present. This study aims to characterize the DNA damage response protein DDR48 and the role that it plays in combating cellular stressors in H. capsulatum. We found that DDR48 is expressed strongly in the mold phase but expressed only …

Cloning, Expression, And Purification Of Fadk And Its Application In Coa-Rna Capture, Jon-Michael L. Stork

Honors Theses

Coenzyme A (CoA) is an important enzyme cofactor involved in acyl transfer reactions. Recently, CoA and its various thioesters were found to exist at the 5’ end of RNA molecules. Although the function of these molecules is still unknown due to difficulties in their isolation, their existence at the 5’ RNA end reveals potentially novel biological roles of RNA. We are exploiting the broad substrate tolerance of the medium chain fatty acid-coenzyme A ligase (FadK), hypothesizing that this enzyme will accept biotinylated medium chain fatty acid substrates, creating biotin-tagged CoA-RNA. This modified CoA-RNA can later be captured using streptavidin affinity …

Signaling Pathways In Skeletal Development And Metabolism, Richard Charles Lindsey

Loma Linda University Electronic Theses, Dissertations & Projects

Osteoporosis, a disease characterized by low bone mass and microarchitectural deterioration, causes nine million debilitating fractures annually. An increase in bone resorption not adequately compensated by a corresponding increase in bone formation is the major cause of osteoporosis. Since current therapies have failed to effectively eliminate osteoporosis, there is an urgent need to discover novel mechanisms regulating bone resorption and formation and develop new therapeutic approaches to treat this disease. To that end, we have explored novel, non-canonical signaling mechanisms of two well-established regulators of bone: thyroid hormone (TH) and vitamin C (ascorbic acid, AA). Since previous studies have shown …

The Effects Of Gibberellic Acid And Auxin Hormones On Heliotropism In Sunflowers, Brandon Bernardo, Hagop S. Atamian

Student Scholar Symposium Abstracts and Posters

Sunflowers are one of many different plant species that are able to track and face the sun in order to optimize the amount of sunlight they are exposed to. This process of orienting towards the sun is called Heliotropism. Sunflowers are able to effectively orient themselves towards the sun because the growth rate on the East and West side of the stem alternates depending on the time of day. At dawn, the East facing stem will grow at a faster rate than the West facing side, resulting in the flower orienting towards the West. This alternating and uneven growth is …

Glycine Receptor Expression Across Identified Retinal Ganglion Cell Types., Ian Scot Pyle

Electronic Theses and Dissertations

Retinal ganglion cells (RGCs) represent the culmination of all retinal signaling and their output forms the substrate for vision throughout the rest of the brain. About 40 different RGC types have been defined by differences in their visually evoked responses, morphology, and genetic makeup. These responses arise from interactions between inhibition and excitation throughout the retinal circuit (Franke et al., 2017; Masland, 2012; Sanes & Masland, 2015; Werblin, 2011). Unlike most other areas of the central nervous system (CNS), the retina utilizes both GABA and glycine inhibitory neurotransmitters to refine glutamatergic excitatory signals (Franke & Baden, 2017; Werblin, 2011; C. …

The Q Rule In Bacteriodetes And The Identification And Characterization Of Porphyromonas Gingivalis Glutaminyl Cyclase., John Andrew Houston

Electronic Theses and Dissertations

Background: Porphyromonas gingivalis, a major pathogen associated with chronic periodontitis, secretes variety of proteins, majority of which begins with glutamine. Several of these proteins were found with pyroglutamate (pGlu) at N-terminus suggesting the presence of this posttranslational modification pathway in P.gingivalis. The observation that N-terminal glutamine is over-represented as the first amino acid after signal peptide cleavage, and subsequent confirmation of pGlu formation on the nascent protein via mass spectrometry, led us to conclude that an enzyme must be present as the executor of this reaction. Hypothesis: PG2157 is a glutaminyl cyclase and is responsible for the cyclization of …

Development Of A Lectin-Fc Fusion Protein With Antiviral And Anti-Cancer Activity., Matthew William Dent

Electronic Theses and Dissertations

This thesis describes the development of a novel lectin-Fc fusion protein and its antiviral and anti-cancer activity. The molecule, Avaren-Fc (AvFc), is a fusion of a variant of the actinomycete lectin actinohivin (Avaren) and the Fc region of human IgG1, and is selective for the terminal α1,2-mannose residues found at the ends of high-mannose-type glycans that can be found on the surface of certain heavily glycosylated viruses and cancer cells. Here, AvFc was found to be able to neutralize simian immunodeficiency virus as well as Hepatitis C virus with nanomolar IC₅₀ values. Furthermore, AvFc recognizes a number of cell …

Paraoxonase 2 Is Critical For Non-Small Cell Lung Carcinoma Proliferation., Aaron Whitt

Electronic Theses and Dissertations

Non-small cell lung carcinoma (NSCLC) comprises 85% of lung cancer diagnoses and is plagued by drug resistance. Thus, elucidating the underlying mechanisms of NSCLC is paramount to expand future treatment options. Paraoxonase 2 (PON2), an intracellular enzyme with arylesterase and lactonase functions, has well-established anti-atherosclerotic activity. Recent studies show PON2 is overexpressed in a variety of tumors and confers drug resistance, although these interactions have not been thoroughly examined in NSCLC. Thus, we sought to investigate the role of PON2 in cellular proliferation using PON2-knockout mice, primary mouse cells, and NSCLC cell lines. Using these approaches, we demonstrate that PON2 …

An Oxanthroquinone Derivative Disrupts Ras Plasma Membrane Localization And Function By Inhibition Of Acylpeptide Hydrolase And Perturbation Of Sphingomyelin Metabolism, Lingxiao Tan

Dissertations and Theses (Open Access)

Oncogenic RAS proteins are commonly expressed in human cancer. To be functional, RAS proteins must undergo post-translational modification and localize to the plasma membrane (PM). Therefore, compounds that prevent RAS PM targeting have potential as putative RAS inhibitors. Here we examined the mechanism of action of oxanthroquinone G01 (G01), a recently described inhibitor of KRAS PM localization. We show that G01 mislocalized HRAS and KRAS from the PM with similar potency and disrupted the spatial organization of RAS proteins remaining on the PM. G01 also inhibited recycling of epidermal growth factor receptor and transferrin receptor, but did not impair internalization …

Spatial Reorganization Of Histone-Like Nucleoid Structuring Proteins Caused By Silver Nanoparticles, Meaad Alqahtany

Graduate Theses and Dissertations

Silver nanoparticles (AgNPs) and ions (Ag+) can be the new generation of antibiotics due to their antimicrobial effects against bacteria and other microbes. Many studies have shown that AgNPs and suppress the growth of bacteria and damage the cell walls of the microbes; therefore, treating bacterial cells with AgNPs may be a promising method to terminate multi-resistant bacteria. In this work, the effect of AgNPs with two different surface coatings on the spatial reorganization of histone-like nucleoid structuring (H-NS) proteins in Escherichia coli bacteria was investigated using quantitative super-resolution fluorescence microscopy to understand the toxicity and antimicrobial mechanism of AgNPs. …

The Role And Regulation Of Alternative Polyadenylation In The Dna Damage Response, Michael R. Murphy

Dissertations, Theses, and Capstone Projects

Cellular homeostasis is achieved by the dynamic flux in gene expression. Post-transcriptional regulation of coding and non-coding RNA offers a fast method of adapting to a changing cellular environment, including deadenylation, microRNA (miRNA) pathway, and alternative polyadenylation (APA). In this dissertation, I explored some of the mechanisms involved in the post-transcriptional regulation of gene expression. The main hypothesis in these studies is that a single APA event after DNA damage is governed by specific conditions and factors outside of current known regulators of APA, and that the resultant transcript has a role in the DNA damage response (DDR). My aims …

Characterization Of Extracellular Vesicles From Human Diabetic Retinopathy Retinal Tissue In Vitro And From Urine Of Human Patients With Diabetic Retinopathy, Jason Mighty

Dissertations, Theses, and Capstone Projects

Diabetic Retinopathy (DR) is a neurovascular complication associated with diabetes mellitus that affects approximately 120 million people worldwide and its prevalence is expected to reach 190 million by 2030. DR diagnosis is accomplished with fundus ophthalmoscopy often when retinal damage and vision loss have already occurred. A group of biomarker being explored for early detection of diseases are extracellular vesicles (EVs), which are nanometer diameter lipid enclosed vesicles, released from all cell types and containing genetic cargo reflective of releasing cell state. EV biomarkers are currently being explored to help monitor disease predisposition, pathogenesis and response to treatment. While an …

The Role Of Parathyroid Hormone 1 Receptor (Pth1r) Signaling In Gastric Epithelial Homeostasis, Maram Maher Al-Hasan

Theses

In the stomach, the epithelial stem cells are responsible for glandular homeostasis. These stem cells continuously undergo cellular proliferation and differentiation to balance the death of senescent cells. Studies conducted on gastric cancers showed a significant increase in parathyroid hormone-like hormone (PTHLH). Moreover, both PTHLH along with parathyroid hormone (PTH) act as endogenous ligands for parathyroid hormone 1 receptor (PTH1R) which belongs to family B of G- protein-coupled receptors (GPCRs). This receptor functions by mediating many different signaling pathways and the most studied pathway is the stimulation of cyclic adenosine monophosphate (cAMP) as a second messenger.

There are very few …

An Investigation Into Clinically Relevant Determinants Of Azole Resistance In Candida Albicans, Andrew T. Nishimoto

Theses and Dissertations (ETD)

"Candida albicans is a commensal organism commonly colonizing the human gut and skin. As an opportunistic pathogen, it can cause persistent and serious infections in individuals with compromised immune systems, including the very young and elderly. Moreover, C. albicans can cause a wide spectrum of diseases ranging from superficial mucosal infections to life-threatening invasions of the organs and bloodstream. Candida species are the most common cause of invasive fungal disease, which is associated with high mortality and imposes a heavy toll on the healthcare system. Over the last 30 years, the azole antifungals have been a mainstay of antifungal therapy, …

Thiol-Based Misfolding: Linking Redox Balance To Cytosolic Proteostasis, Ford Amy

Dissertations and Theses (Open Access)

The eukaryotic cytosolic proteome is vulnerable to changes in proteostatic and redox balance caused by temperature, pH, oxidants and xenobiotics. Cysteine-containing proteins are especially at risk as the thiol side chain is subject to oxidation, adduction and chelation by thiol-reactive compounds. All of these thiol-modifiers have been demonstrated to induce the heat shock response and recruit protein chaperones to sites of presumed protein aggregation in the budding yeast Saccharomyces cerevisiae. However, endogenous targets of thiol stress toxicity responsible for these outcomes are largely unknown. Furthermore, I hypothesize proteins identified as redox-active are prone to misfolding and aggregation by thiol-specific …

Hsp70-Mediated Regulation Of Hsf1 Transcriptional Activity In Saccharomyces Cerevisiae, Sara Peffer

Dissertations and Theses (Open Access)

In eukaryotic cells, protein homeostasis and cellular fitness is promoted by the transcription factor heat shock factor 1 (HSF1) during exposure to proteotoxic stress. HSF1 controls the basal and stress-induced expression of molecular chaperones and other protective targets. Dynamic regulation of HSF1 involves the major heat shock proteins Hsp70 and Hsp90. Recent advances in the understanding of this regulatory circuit in Saccharomyces cerevisiae have shown that the Hsp70 Ssa1 acts as a sensor for some proteotoxic stresses and is capable of a direct interaction with Hsf1. This work continues to explore the complex regulatory interaction between Hsf1 and Ssa1. I …

The Role Of Gene Expression Noise In Mammalian Cell Survival, Kevin Farquhar

Dissertations and Theses (Open Access)

Drug resistance and metastasis remain obstacles to effective cancer treatment. A major challenge contributing to this problem is cellular heterogeneity. Even in the same environment, cells with identical genomes can display cell-to-cell differences in gene expression, also known as gene expression noise. Gene expression noise can vary in magnitude in a population or in fluctuation time scales, which is influenced by gene regulatory networks.

Currently, it is unclear how gene expression noise from gene regulatory networks contributes to drug survival outcomes in mammalian cells. An isogenic cell line with a noise-modulating genetic system tuned to the same mean is required. …

Development Of A High-Throughput System For Screening Of Anti-Prion Molecules, Katherine Do

Dissertations and Theses (Open Access)

The misfolded prion protein causes and transmits disease in both humans and animals. As other infectious agents, prions display strain variation, which can generate different pathological outcomes in affected individuals. Unfortunately, there are no known therapies for these diseases, which at present are invariably fatal. In this work, the Protein Misfolding Cyclic Amplification technology (PMCA, an in vitro test that replicates minimum quantities of infectious prions) has been modified to screen for small molecules inhibiting prion protein misfolding in a strain-specific manner. In order to approach a high-throughput PMCA system, technical aspects in PMCA has been optimized for application of …

Preparation Of A Flavonol Specific Glucosyltransferase Found In Grapefruit And Site-Directed Mutants For Protein Crystallization, Aaron Birchfield

Electronic Theses and Dissertations

This research was designed to determine the conditions necessary to remove c-myc and 6x-His tags from a flavonol specific glucosyltransferase found in grapefruit (CP3GT) using thrombin in preparation for crystallization. X-ray crystallography of CP3GT crystals may elucidate structural features that account for flavonol specificity in some glucosyltransferase enzymes. A thrombin cleavage site was inserted into WT CP3GT and one mutant. Recombinant CP3GT was expressed in yeast and purified. Optimal conditions for thrombin digestion were explored. Digestion with 100U of thrombin for 2 hours at 4^o C was optimal for removing tags from CP3GT. Storage at 4^o C for …

Hormone Signaling, Gene Expression, And Mitochondrial Hormone Receptor Expression In Avian Muscle (Cells), Kentu Rushadd Lassiter

Graduate Theses and Dissertations

Mitochondria are vital to the proper growth and function of muscle cells since they’re responsible for the majority of ATP production used for cellular energy. Previous studies have investigated how differences in mitochondrial function affects feed efficiency (FE) in broilers phenotyped for High and Low FE. Low FE broilers have been shown to have increased levels of reactive oxygen species (ROS), thus contributing to higher levels of oxidative stress and damage seen in these birds. Global gene and protein expression studies conducted on breast muscle of the High FE and Low FE phenotypes have suggested that differences in mitochondrial function …