Analytical Chemistry Commons^™

Amyloid Fibril Formation And Polymorphism : A Critical Role Of Sulfur-Containing Amino Acid Residues, Tatiana Quiñones-Ruiz

Legacy Theses & Dissertations (2009 - 2024)

Protein aggregation that results in the formation of amyloid fibrils has been linked to many neurodegenerative disorders, including Alzheimer’s disease and Parkinson’s disease. The sulfur atoms in methionine (Met) and cysteine (Cys) residues of proteins can be readily oxidized, significantly affecting their properties. Oxidation of sulfur-containing amino acids has recently been shown to affect protein fibrillation. This work presents novel findings on Cys and Met redox reactions that are related to the formation of amyloid fibrils and on the polymorphism of a model fibrillogenic protein, hen egg white lysozyme (HEWL). Biophysical techniques including Raman spectroscopy, atomic force microscopy, electron paramagnetic …

An Ims-Ms/Md Workflow For Determining Higher Order Structure And Dynamics Of Nucleic Acids, Rebecca D'Esposito

Legacy Theses & Dissertations (2009 - 2024)

Ion mobility spectrometry - mass spectrometry (IMS-MS) has potential for the investigation of structure and dynamics in large biopolymers, which will come to full fruition only with a firmer understanding of how to interpret the experimental data. Numerous studies have employed elements of nucleic acid (NA) secondary structure, such as duplexes and hairpins, to explore the relationships between structure, experimental conditions, and actual observations. When combined with molecular dynamics simulations (MDS), IMS-MS can be effectively employed to perform structural elucidation of biomolecules that are not readily amenable to established techniques employed for structural analysis.

An Analytical Method To Detect And Quantify Ll-37 In An In Vitro Co-Culture Of Activated Thp-1 Cells And Staphylococcus Aureus, Staphylococcus Epidermidis, And Pseudomonas Aeruginosa, Victoria Hunter

Graduate Theses and Dissertations

Healthcare-associated infections (HCAIs) affect 1.7 million hospitalized patients each year, resulting in over 98,000 deaths and anywhere from $28.4 to $45 billion in treatment costs. Furthermore, it has been established that more than 80% of these infections are caused by biofilms While HCAIs can be bacterial, viral, fungal, or parasitic, there are between 12 and 17 species cause 80 to 87% of all HCAIs. Among the most commonly isolated microorganisms are Pseudomonas aeruginosa (P. aeruginosa) and Staphylococcus aureus (S. aureus). Thus, biofilms are of importance not only due to their prevalence in HCAIs but also due to the increased antibiotic …

A New Insight Into Fungal Cell Wall Architecture By Functional Genomics And Solid-State Nmr Along With Recent Advancements In Dynamic Nuclear Polarization For Analyzing Biomolecules, Arnab Chakraborty

LSU Master's Theses

This dissertation summarizes the findings related to the way by which supramolecular architecture of fungal cell wall changes with genetic mutation, dispensing genes responsible for biosynthesis of cell wall polysaccharides. This is necessary because without perfect picture of how supramolecular assembly changes with genetic mutation it is hard to assess new anti-fungal targets. Alongside this we have highlighted how recent advancement into Dynamic Nuclear Polarization (DNP) methods improved characterization of biomolecules both in case of labeled and unlabeled samples.

First study utilized Solid-state NMR (SSNMR) which is a non-destructive technique hence enabled us for the first time to deduce how …

Analysis Of The Light Responsive Azobenzene Peptide Nucleic Acid Duplexes, Kat Nguyen

Honors College Theses

Peptide nucleic acids (PNAs) are oligonucleotide analogues in which the sugar-phosphate backbone has been replaced by a pseudopeptide skeleton. Since PNAs use the natural nucleobases (Adenine, Thymine, Cytosine, Uracil, and Guanine) found in either DNA and/or RNA, they are able to hybridize according to Watson-Crick base-pairing to form duplexes. PNA is a promising therapeutic agent because they can function as antigene or antisense chemical agents. To further enhance their utility, we aim to incorporate a photoswitchable moiety using azobenzene. Here, we report the results of the synthesis and purification of a photoswitchable 11 mer PNA along with initial characterization efforts.

Design, Synthesis, And Analysis Of Paired Coiled-Coil Peptidic Molecular Building Blocks Used For Linearly Controlled Self-Assembly Of Α-Helical Coiled-Coil Heterodimer Peptide Pairs, Jason Distefano

Chemistry Theses

Molecular building blocks are fundamental to biological synthesis and processes and have been utilized in advanced materials, drugs and drug delivery systems, and biotechnology. Proteins have been used as molecular building blocks for the construction of complex, well-ordered structures. Coiled-coil protein domains are essential subunits used for the oligomerization of protein complexes, gene expression, and structural elements of biological materials. The synthesis and assembly of proteins utilizing coiled-coil motifs are of great scientific interest due to their potential applications in disease treatment, biomechanical motors, nanoscale delivery systems, etc. However, assembling protein complexes with specific morphology is still challenging because …

Extending Plasma Proteome Coverage To Enable Biomarker Discovery, Jordan Bruce Burton

Wayne State University Dissertations

Two features make plasma a valued source of health information: First, plasma contains secreted proteins from all tissues in the body and can, therefore, provide information about the health status of all tissues. Second, plasma is available in large quantity using a minimally invasive procedure that is well tolerated. The status of plasma as a valued source of health information is matched by the difficulty in extracting proteome information from the samples. Using state of the art instrumentation a standard analysis of plasma yields quantitative information on approximately 300 proteins. Extreme fractionation strategies coupled with increased mass spectrometry time extends …

Building Tools For Improved Modulation Of The Human Gabaa Receptor, A Central Nervous System Target For The Treatment Of Anxiety, Garrett Edward Zinck

Theses and Dissertations--Pharmacy

In the U.S., anxiety is recognized as an increasing range of mentally and physically debilitating psychiatric health disorders with significant economic repercussions. Over the last 20 years, several novel anti-anxiety therapies have entered the drug development pipeline, but none have made it to market.

The work in this dissertation focused on structurally modifying valerenic acid (VA), a structurally unique carboxylated sesquiterpene acid found in Valeriana officinalis. VA is putatively reported to have allosteric modulatory activity of the human GABA_A receptor, a ligand-gated ion channel responsible for attenuating neurotransmissions. Structural modeling of VA’s GABA_A receptor interaction suggests that …

Purification And Activity Assay Optimization Of Bacterial Catalase From Psychrobacter Koprana, Sydney Willis

Honors Program Theses

To date, catalase protein from bacterial species is not the most widely used commercial catalase. Catalase, the enzyme that breaks down hydrogen peroxide in most living organisms, is an invaluable enzyme in the food, agriculture, and pharmaceutical industries. In 2019, a novel species of a psychrotolerant bacterium was discovered in Cold Harbor, NY. This species, Psychrobacter koprana, exhibits highly active catalase. Preliminary data from activity assays demonstrates that P. koprana catalase has the same activity as commercially available catalase. After purification and activity assay optimization, the activity of this catalase has the potential to be more active than other easily …

Development Of Fluorescence Based Approaches To Understand Astrocyte Biology In The Context Of Nicotine And Nicotinic Receptor Activity, Surya P. Aryal

Theses and Dissertations--Chemistry

Smoking and tobacco use (STU) is a major global health problem and worldwide more than six million people die due to tobacco related diseases each year. Although majority of smokers try to quit smoking several times in their life, traditional therapeutic approaches, which focus only on neuronal cells, have a very low success rate. Understanding the effect of nicotine on glial cells, synaptic communication and blood vasculature in the brain can provide further insights on the neurobiology of substance abuse and can potentially help to design better therapeutic approaches. Glial cells are non-excitable cells in the brain which do not …

Effects Of Localized Oxygen Production By Electrolysis On The First-Generation Glucose Sensor Response, Nandita Halder

Graduate Theses and Dissertations

Glucose sensors are very important for detecting blood glucose both in vitro and in vivo. First-generation glucose biosensors were based on the glucose oxidase (GOx) enzyme using molecular oxygen as the electron acceptor and therefore oxygen dependent. Unfortunately for in-vivo work, oxygen in the body is variable and limited. Alternative approaches to overcome the oxygen dependency came with their own limitations. The widely used and commercially available ex-vivo glucose test strip uses a mediator in place of oxygen to free it from oxygen dependency. The mediator-based technology, in most cases cannot be transferred to in vivo applications due to the …

3-Dimensional Structural Determination Of Epidermal Growth Factor-Like Repeat 27 From Mouse Notch1 Using Nuclear Magnetic Resonance Spectroscopy, Justin Grennell

LSU Doctoral Dissertations

NOTCH1 has been associated with a variety of human ailments, such as cancer and heart disease. NOTCH1 is a transmembrane protein that contains 36 epidermal growth factor-like (EGF) repeats, which facilitate cell signaling by binding proteins on neighboring cells. Each repeat contains about 40 amino acids and has to be folded in a specific manner to function properly. The amino acid sequence, carbohydrate pattern, and calcium-binding sites of EGF-like repeats vary, but the pattern along the 36-repeat chain is conserved. The Abruptex region (EGF24-29) of NOTCH1 is a highly glycosylated (having undergone sugar addition) and flexible region, which gained its …

Don't Sell Them Short, There's More To Bacterial Natural Products Than Antibiotics, Alison Clare Domzalski

Dissertations, Theses, and Capstone Projects

Recent genomic studies of microbiomes have revealed an overwhelming number of biosynthetic genes of unknown function. Most of these “cryptic” biosynthetic genes are not expressed in laboratory monocultures of individual microbes. Thus, there remains tremendous untapped potential for natural products discovery. Here we employ mixed microbial culture (MMC) as a simple yet powerful approach to awaken cryptic biosynthetic gene clusters. Our preliminary studies demonstrated that arrays of metabolites could be induced in MMCs upon environmental cues, such as surface adhesion. Using this system, we have screened, identified, and isolated bioactive bacterial metabolites, which were characterized structurally and biologically. Of the …

Indicators Of Senescence And Their Relationship With The Antioxidant Response Element In Ovarian Cancer Cells—Toward A More Encompassing View Of Enzyme Activity Levels In Cells, Shaniqua Analycia Hayes

LSU Doctoral Dissertations

Enzymes and other related proteins are promising targets as biomarkers for cancer diagnostics, and it is therefore important to be able to quantify intracellular enzyme activity levels, not just presence of enzyme. Thus, there is a great need for the development of methods that offer spatiotemporal quantification of intracellular enzyme activity levels in respiring 2- and 3-dimensional cultures or their sample-processed variants. Previous studies have demonstrated that ß-gal is upregulated in senescence and in select ovarian cancers, and its expression/activity is upregulated when cells are under oxidative stress conditions. To that end, the primary goal of this work was to …

Quantifying Phosphatidylethanol From Blood Samples, James V. Defrancesco, Majid Afshar

Chemistry: Faculty Publications and Other Works

Disclosed herein is a method for quantifying phosphatidylethanol (“ PEth ”), direct ethanol biomarker, from a blood sample using gas chromatography mass spectrometry. The method disclosed herein is useful for diagnosing acute and chronic alcohol abuse.

Bioanalytical Determination Of Glucose Concentration In Sports Drinks Using Uv/Vis Spectroscopy, Laney Hedgeman, Brianna Bond

Symposium of Student Scholars

Abstract

The purpose of our project is to indirectly monitor the enzymatic activity of glucose oxidase as it reacts with the glucose present in sports drinks through the quantitative analysis of the generated ferricyanide. As glucose oxidase catalyzes the oxidation of beta-D-glucose in the presence of oxygen, D-glucono-1,5-lactone is produced along with hydrogen peroxide. The hydrogen peroxide is used in a subsequent reaction with ferrocyanide catalyzed by horse radish peroxidase to produce water and the chromophore ferricyanide. This compound absorbs in the ultraviolet/visible spectrum at 420 nm, which can be quickly measured using Ultraviolet/Visible Spectroscopy. Since the molar ratio of …

Investigations Of The Structure And Protein-Protein Interactions Of Chlamydia Trachomatis Scc4, Thilini Oshadhi Senarath Ukwaththage

LSU Doctoral Dissertations

Chlamydia trachomatis (CT) is the most common, sexually transmitted bacterial disease (STD) in the world. In the developmental cycle of CT, specific chlamydia chaperone 4 (Scc4) is a unique protein with essential and multiple roles. Hence, Scc4 is significant as a virulence target for therapeutic approaches to treat chlamydial infections. A novel approach was discovered to purify tag free Scc4 by utilizing a 6X-histidine-tag on Scc1 in the co-expressed Scc4:Scc1 complex by capturing the complex on nickel-charged immobilized metal affinity chromatography resin, followed by dissociation of Scc4 with sarkosyl. Using triple resonance NMR experiments, backbone and sidechain resonances …

Using The Marcus Inverted Region And Artificial Cofactors To Create A Charge Separated State In De Novo Designed Proteins, Eskil Me Andersen

Dissertations, Theses, and Capstone Projects

To create an efficient de novo photosynthetic protein it is important to create long lived charge separated states. Achieving stable charge separation leads to an increase in the efficiency of the photosynthetic reaction which in turn leads to higher yields of end products, such as biofuels, electrical charge, or synthetic chemicals. In an attempt to create charge separated states in de novo proteins we hypothesized that we could engineer the free energy gaps in the proteins from excited primary donor (PD) to acceptor (A), and A back to ground state PD such that the forward electron transfer (ET) would be …

Validation Of A Deployable Proteomic Assay For The Serological Screening Of Sexual Assault Samples, Catherine O'Sullivan Brown

Electronic Theses and Dissertations

Protein mass spectrometry (MS) has emerged as a technique to supplant traditional serological tests for body fluid identification. It was hypothesized that proteomic techniques would surpass the sensitivity and specificity of traditional serological techniques. An automated workflow coupled with protein MS has been developed for the confirmatory identification of five biological fluids. A developmental validation was completed, assessing parameters such as reproducibility, sensitivity, ion suppression, and limit of detection. Implementation was determined through tandem sample processing by MS, traditional serological tests, and standard DNA profiling methods. The MS approach offered superior detection limits while also providing true confirmatory results, producing …

High Throughput Analysis To Study Emerging Pollutants And Nanoparticle Fate In Biological Systems, Xiaolong He

Doctoral Dissertations

”The increasing applications of emerging and fugitive contaminants (EFCs) and engineered nanoparticles (ENPs) attract significant research interest for their potential risks to human health and the environment. In order to assess the health risks of these emerging contaminants, rapid and reliable analytical methods to measure the concentrations and fates of these contaminants are imperative. This dissertation focuses on the developments of advanced analytical methods and their applications to study those emerging contaminants in crop plant and simulated gastric fluid (SGF). Three types of mass spectrometry based methodologies have been developed, one is freeze-thaw/centrifugation extraction followed by high performance liquid chromatography …

The Quantitative Assessment Of Pond Scum: An Examination Of The Biogeochemistry Of Phosphorus Cycling In The Belgrade Lakes, Abbey M. Sykes

Honors Theses

The internal recycling phosphorus in freshwater lake bottom sediments represents a significant source of hypolimnetic phosphorus (P) release for many of Maine’s lakes. In summer months, Maine lakes often thermally stratify and the lake hypolimnion develops anoxia, leading to a reduction in redox potential at the sediment-water interface. These reducing conditions facilitate the reductive dissolution of ferric iron, and, since phosphorus is often present in freshwater lake sediments as solid FeOOH-PO₄ complexes, results in release of soluble phosphorus into the water column. Our current study presents field and laboratory data from sediment fractionation extractions designed to quantify concentrations of …

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 …

Using Nmr Spectroscopy And Computational Chemistry To Confirm The Structure Of Novel Antibiotic Nocamycin O, Stephanie Lewis

CMC Senior Theses

In recent years, many medically promising antibiotics have been discovered in nature, especially in insect-microbe symbioses. One of the better-studied examples of this kind of defensive relationship is that of fungus-growing ants and the antibiotic-producing Actinobacteria. These bacteria produce several defensive chemicals with myriad uses, including one antibiotic that inhibits the growth of several bacterial strains, including other Actinobacteria. This antibiotic (known as nocamycin O) is a promising candidate for medicinal use due to its similarities to bacterial RNA polymerase inhibitors tirandamycin and streptolydigin, which inhibit several human pathogens. The determination of the structure of nocamycin O will be an …

Flavin Modification And Redox Tuning In The Bifurcating Electron Transfer Flavoprotein From Rhodopseudomonas Palustris: Two Arginines With Different Roles, Nishya Mohamed-Raseek

Theses and Dissertations--Chemistry

Electron bifurcation is considered as a third fundamental mode of energy conservation mechanism in addition to two well-known mechanisms, substrate level phosphorylation and Oxidative phosphorylation, in electron bifurcation endergonic and exergonic redox reactions are coupled. The newly discovered flavin based electron bifurcation in electron transfer flavoproteins (ETFs) helps to reduce low potential ferredoxin, which provides electrons to drive biologically demanding reactions such as atmospheric dinitrogen fixation in diazotroph and methane production in methanogens.

Current research demonstrates the capacity for electron bifurcation in the Rhodopseudomonas palustris ETF (RpalETF) system. RpalETF contains two chemically identical but functionally different FADs: …

Graphene-Based Electrochemical Aptasensor For The Detection Of Salmonella Typhimurium, Shalini Muniandy

Student Works (2020-2029)

Recent foodborne outbreaks in multiple locations necessitate the continuous development of highly sensitive and specific biosensors that offer rapid and sensitive detection of foodborne pathogens. The emergence of electrochemical aptasensor which is a compact analytical device with a combination of an aptamer as the biorecognition element integrated on the transducer surface created a breakthrough in the detection of foodborne bacterial pathogens. Graphene has emerged as a promising nanomaterial for reliable detection of pathogenic bacteria due to its exceptional properties such as high electrical conductivity, large surface to volume ratio, high mechanical strength and its unique interactions with DNA bases of …

Application Of Carbon Nanoparticles As Dna Detection Probe And Fluorescent Ink, Luckio Frank Owuocha

Graduate Theses/Dissertations

There is a significant interest in developing a sensitive, selective, efficient, and inexpensive method for rapid molecular diagnostic tests. This research aims to develop an inexpensive nucleic acid detection method by using DNA-conjugated carbon nanoparticles that exhibit fluorescence in the visible region. Carbon nanoparticles of this class can be detected without specialized equipment and have great promise toward the development of analytical methods that can be used in resource-limited environments with a lack of access to proper diagnostic and healthcare. We employed EDC-NHS (two-step) and EDC (one-step) coupling techniques to prepare DNA-conjugated carbon nanoparticles. The dot blotting method was adapted …

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 …

Intrinsic Buffer Hydroxyl Radical Dosimetry For Hydroxyl Radical Protein Footprinting, Addison Roush

Honors Theses

Hydroxyl radical protein footprinting (HRPF) coupled to mass spectrometry is a powerful technique for the analysis of protein topography as it generates covalent mass labels that can survive downstream sample handling, and it is sensitive to the solvent accessibility of amino acid sidechains. Of the multiple platforms for HRPF, fast photochemical oxidation of proteins (FPOP) utilizes a pulsed 248 nm KrF excimer laser to label proteins by photolyzing hydrogen peroxide. FPOP is the most widely used HRPF platform because it labels proteins faster than unfolding can occur. Variations in FPOP sample conditions make it difficult to compare results between experiments …

Assessment Of Soil Protein And Refractory Soil Organic Matter Across Two Chronosequences Of Newly Developing Marshes In Coastal Louisiana, Usa, Stuart Alexander Mcclellan

LSU Doctoral Dissertations

The impacts of sea-level rise and hydrologic manipulation are threatening the stability of coastal marshes throughout the world, thereby increasing the potential for re-mineralization of soil organic matter (SOM) in these systems. Such threats have prompted marsh restoration efforts, particularly in coastal Louisiana, yet it is unclear how the slowly decomposing (refractory) and quickly decomposing (labile) fractions of SOM may be differentially affected by different approaches to marsh restoration. Additionally, otherwise labile compounds may accumulate in the soil via a range of protective mechanisms, including rapid burial and association with organic compounds that are thought to enhance soil aggregation, such …

Yeast Copper Proteins And Reactive Oxygen Species In Effecting Lifespan, Megan Bestwick

Post-Grant Reports

Mitochondria are essential organelles in most eukaryotic cells because of their role in metabolism and the production of ATP by the oxidative phosphorylation (OXPHOS) pathway, as well as other key cellular processes. Metal cofactors, such as copper (Cu) and iron (Fe), are incorporated into OXPHOS protein complexes of yeast located within the inner membrane of the mitochondria. Misincorporation or modulation of these available metals in mitochondrial enzymes leads to the production of reactive oxygen species (ROS). ROS are reactive molecules containing oxygen such as peroxides, superoxide, and hydroxyl radicals. Yeast are a good model for studying aging and the effect …