Physical Sciences and Mathematics Commons^™

Targeting Heat Shock 27 Kda Protein Induces Androgen Receptor Degradation, Yaxin Li

ETD Archive

Glioblastoma (GBM) is the most common and aggressive brain tumor, with very poor prognosis. Androgen receptor (AR) plays a significant role in the progression of GBM, and anti-androgen agents have the potential to be used for the treatment of GBM. However, AR mutation commonly happens in GBM, which makes the anti-androgen agents less effective. Heat shock 27 kDa protein (HSP27) is a well-documented chaperone protein to stabilize AR. Inhibition of HSP27 results in AR degradation regardless the mutation status of AR, which makes HSP27 a good target to abolish AR in GBM. Identified compound I ((N-(3-((2,5-dimethoxybenzyl)oxy)-4-(methylsulfonamido) phenyl)-4-methoxybenzamide) inhibits GBM cell …

Covalent Modification Of Recombinant Protein With Reactive Thiols, Sawyer Dulaney, Bailey Taylor

Honors Theses

Many diseases cause chronic and painful inflammation in different body systems. One of the front-line drug classes to treat such inflammation is Nonsteroidal Anti-Inflammatory Drugs (NSAIDs). Despite the benefits of oral administration of NSAIDs, there are drawbacks to their long-term usage because they can cause detrimental effects on off-target systems in the body such as the liver, kidney, or the lining of the intestinal tract. An alternative to NSAIDs is the usage of hydrogels for targeted drug delivery. Hydrogels can provide drug delivery in a specific portion of the site of inflammation, thus allowing higher doses of medication to be …

Biochemical Characterization Of Induced Pluripotent Stem Cell-Derived Cardiomyocytes As A Model Of Barth Syndrome, Alisha J. House

ETD Archive

Barth Syndrome (BTHS) is an X-linked inborn error of metabolism (IEM) which manifests as a multi-systemic disease. One of the primary symptoms is dilated cardiomyopathy, and alongside the cardiovascular disease that arises, patients often experience metabolic abnormalities such as 3-methylglutaconic aciduria, growth retardation, and neutropenia. There has been a need for the development of a suitable in vitro modeling system which will accurately recapitulate the biochemical and physical nature of BTHS. The purpose of this project has been to develop a model for studying the biochemical pathogenesis of Barth Syndrome using human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). To achieve …

Investigating Spatiotemporal Kinetics, Dynamics, And Mechanism Of Exosome Release, Anarkali Mahmood

Electronic Theses and Dissertations

Exosomes are small lipid-based vesicles that can carry biomolecules from one cell to another. While exosomes are crucial to maintain homeostasis in healthy cells, they are exploited by unhealthy cells to aid disease progression. Exosomes likely facilitate disease progression via the transfer of disease-causing biomolecules from unhealthy to healthy cells. Exosomes are generated in Multivesicular endosomes (MVEs) and are then secreted into the extracellular space to travel to other cells. Despite being a crucial step, very little is known about exosomes release mechanism and dynamics. To further our understanding of exosomes, specifically their secretion, my work has focused on investigating …

Inhibitors Of Alpha-Synuclein Aggregation, Jemil Ahmed

Electronic Theses and Dissertations

Alpha-Synuclein (αS) – a neuronal, disordered, presynaptic protein – aggregates into amyloid fibrils and accumulates in the substantia nigra pars compacta of Parkinson's Disease (PD) patients. The aggregation and accumulation of αS amyloid fibrils leads to death of dopaminergic neurons; a hallmark of PD. Although it’s not clear why αS aggregates, prior studies have found that intrastriatal injection of fibril alone is sufficient to cause PD pathology in mouse and non-human primates models. These observations implicate αS as a therapeutic target against PD.

Unfortunately, there are three caveats when attempting to target αS. First, αS is a neuronal protein expressed …

Structural Characterization Of Factor Viii-Inhibitor Complexes And Factor Viii Lipid Binding Mechanics, Corbin Mitchell

WWU Graduate School Collection

Blood coagulation factor VIII (FVIII) is a crucial protein cofactor within the blood coagulation cascade and facilitates the proteolytic activation of factor X by activated factor IX. During coagulation FVIII is activated and binds, via its C1 and C2 domains, to activated platelet membranes coordinated by interactions with exposed phosphatidylserine on the membrane surface. A deficiency of functional FVIII within a patient's bloodstream leads to the blood disorder hemophilia A, which results in prolonged bleeding episodes. Current treatment for hemophilia A relies on FVIII replacement therapy via the injection of exogenous FVIII. The main complication which arises from FVIII replacement …

Ahr Expression On Rorc-Expressing Immune Cells Is Essential For I3c-Mediated Protection Against Colitis, Michal C. Williams

Senior Theses

Colitis is an inflammatory bowel disorder (IBD) whose etiology is attributed to modification in the luminal microbiota and dysregulation in the immune response. Indole is a signaling molecule which is naturally produced by gut luminal microbiota. Indole-3- carbinol (I3C) is a compound commonly found in vegetables and a ligand for the aryl hydrocarbon receptor (AhR). Previous studies have detected decreased expression and activation on the AhR receptor in colitis patients, thought to possibly alter gut microbiota metabolism, subsequently promoting colitis. 1 AhR, expressed in a variety of immune and epithelial cells, contributes to gut homeostasis by affecting vital mediators such …

Effects Of Crowding Agents On I-Motif Dna, Hayden Brines

Honors Theses

Deoxyribonucleic acid (DNA) is a well-known double stranded, helical, biological molecule. In addition to its more commonly known structure, DNA can also form more complicated structures like G-quadruplexes and i-motifs (iM). The iMs are formed by cytosine rich DNA and are a four stranded structure that is typically looped around itself. The iM formation is typically pH-dependent and is favored in more acidic conditions; the pKa value is approximately 6.5. This pKa value allows for potential in vivo formation, since the cells have a pH of approximately 7.3. Due to this, iMs are thought to be powerful, innovative molecules for …

Pectin And Alginate Extraction To Treat Liquid Cafo Manure, Clare Sunderman

Honors Projects

For this project, various extraction methods were used to extract pectin from Pastinaca Sativa and alginate from Macrocystis. These extractions were then dried and used in treating 250mL of manure along with a CaCl₂ or FeCl₃ coagulant. It was found that CaCl₂ was not as effective as FeCl₃ in coagulating manure. But the results obtained suggest that pectin and alginate obtained with a simpler extraction method is just as effective as the highly purified and refined pectin and alginate produced for the food industry, in the treatment of CAFO manure. The liquid portion of the …

Structural And Mutational Characterization Of The Blood Coagulation Factor Viii C Domain Lipid Binding Interface, Shaun C. Peters

WWU Graduate School Collection

Blood coagulation factor VIII (fVIII) functions as a cofactor in the blood coagulation cascade for proteolytic activation of factor X by factor IXa. During coagulation, fVIII is activated and subsequently binds to activated platelet surfaces by coordination of the fVIII C1 and C2 domains to the exposed phosphatidylserine of activated platelet membranes. Structural and mutational studies have suggested that both hydrophobic and electrostatic interactions occur between the two tandem C domains and activated lipid surfaces, but models of C domain phospholipid binding propose conflicting regions that directly interact with the membrane surface. This thesis reports the determination of the molecular …

Methodology For C-Glycosylation And Synthesis Of Probes For The Detection Of Cysteine Containing Proteins, Elizabeth Ward

Undergraduate Honors Theses

Throughout my research I have been able to expand my knowledge and gain experience in organic chemistry by working on projects related to biochemistry and hydrocarbons. The projects I have worked on were:

• Project 1: C-glycosylation through photo irradiation with reductive halide atom transfer
• Project 2: Synthesis of rhodamine and biotin probes for the detection of cysteine containing proteins
• Project 3: Literature review on: Fluorescent probes for cysteine containing proteins

Characterization Of O-Linked Glycosylated Neuropeptides In The American Lobster (Homarus Americanus): The Use Of Peptide Labeling Following Beta Elimination, Edward Myron Bull

Honors Projects

Neuropeptides are a class of small peptides that govern various neurological functions, and the American lobster (Homarus americanus) provides a model system for their characterization. Neuropeptides are commonly post-translationally modified (PTM), and one common PTM is glycosylation. Past research in the Stemmler lab has found glycosylated neuropeptides in H. americanus; however, the extent and biological role of this modification has not been well characterized. This study was undertaken to determine the number of glycosylated peptides in the sinus glands of H. americanus and to develop an approach to tag the site of glycosylation using beta-elimination chemistry. LC-MS …

Characterization Of The Dimerization Domains On The Mannose-6-Phosphate/Insulin-Like Growth Factor Ii Receptor, Tyler Degener

Theses/Capstones/Creative Projects

The mannose-6-phosphate/insulin-like growth factor II (M6P/IGF2) receptor is a transmembrane protein known to sequester growth factors from the extracellular matrix. This behavior suggests a mechanism of tumor suppression. Structurally, the receptor’s extracellular region is segmented into 15 homologous repeats, which are divided further into 5 triplet domains, labelled 1-3, 4-6, 7-9, 10-12, and 13-15. What is notable about the triplets is their propensity to form dimers with triplets on a second M6P/IGF2 receptor. In fact, previous studies indicate that this protein functions optimally when dimerized. Thus, the purpose of this experiment is to characterize these domain interactions. Using a urea …

Evaluating Methods Of Obtaining Male Pheromone From Hymenochirus Sp. Using Analytical Chemistry, Vincent Wing-Kun Leung

University of the Pacific Theses and Dissertations

Male Hymenochirus sp. frogs are known to release pheromone that attracts females of the same species. Four methods for collecting secretions containing pheromone in Hymenochirus sp. were tested: norepinephrine injection, gonadotropin-releasing hormone injection, homogenization of gland tissue, and electrostimulation of the skin over the breeding gland area. The samples collected were analyzed using high-performance liquid chromatography (HPLC) and mass spectrometry. The HPLC chromatograph for the male norepinephrine sample contained a peak at 6.4 min that was not in the female norepinephrine sample HPLC chromatograph. The male norepinephrine sample mass spectrum had a peak of m/z 292.0 not in the female …

Synthesis And Conformational Studies Of Various Amides, Marcos Beltran-Sanchez

University of the Pacific Theses and Dissertations

In the past, aminocyclohexanol rings have been successfully utilized as pH-triggered molecular switches in various trans-2-aminocyclohexanol derivatives. By changing the groups on the amine nitrogen, these models provided a wide pH range in which a switch can occur. The pH-induced switch of conformation was monitored by 1H-NMR spectroscopy. The models were also incorporated into the bilayer membrane of liposome structures and tested for their ability to disrupt their membrane upon their conformational flip induced by a decrease in pH.

In this work, the amide bond has been studied as a molecular switch and various amide derivatives have been tested for …

Use Of Biometrics To Determine Biochemistry Knowledge Level Differences In Reading And Processing Metabolic Pathways, Kim Kammerdiener

Master of Science in Chemical Sciences Theses

The areas of chemistry and biochemistry commonly use external representations to enhance learning for students. The use of schematics and representations for illustrating metabolic pathways is a familiar image in most biochemistry textbooks. These external representations can be the key to student understanding. To date, there has not been any previous research specifically focused on the areas of interest that are predominant for an individual viewing a metabolic pathway commonly found in a biochemistry textbook. Therefore, this thesis study set out to 1) investigate what individuals look at in a metabolic pathway based on their level of expertise in metabolism …

Quantitative Analysis Of Bleomycin In Rat Plasma By Lc-Ms/Ms, Huawen Li

ETD Archive

Bleomycin is the most commonly used compound in its group of antineoplastic drugs. It works on tumor cells by single and double stranded DNA cleavage after its activation, in which it blocks tumor cells’ DNA replication or transcription activities to inhibit tumor cells’ growth. Bleomycin sulfate (Blenoxane) is the most popular preparation used in clinical research, and contains Bleomycin fractions of A2 and B2, which causes difficulties in quantitative analysis. This work uses the metal chelating property of Bleomycin as an advantage to simplify and improve sensitivity of existing quantitative methods. Copper was spiked in excess to plasma samples, followed …

Computational Investigation Of Protein Assemblies, Sm Bargeen Alam Turzo

ETD Archive

Selective nitrosylation of glyceraldehyde 3-phosphate dehydrogenase (GAPDH) at Cys-247 affects gene regulation through the interferon-gamma (IFN- γ) activated inhibitor of translation (GAIT) complex. Oxidized low-density lipoprotein (LDL_ox) and INF-γ induce assembly of the nitrosylase complex composed of inducible nitric oxide synthase (iNOS), S100A8, and S100A9 proteins. Crystal structure of the complex of GAPDH and S100A8A9 is not known, structural prediction method were employed by protein-protein docking and binding energy calculation with PatchDock and FIREDock respectively. Candidate models were selected, based on a weight factor calculated, from the computational method developed from the "artificial protease" cleavage mapping Fe(III) (s)-1-(p- …

Il-17 Drives Copper Uptake And Activation Of Growth Pathways In Colorectal Cancer Cells In A Steap4-Dependent Manner, Evan Martin

ETD Archive

Colorectal cancer is a disease characterized by abnormal, invasive cell growth beginning in the colon or rectum. The third most common type of cancer worldwide, approximately one million new cases of the disease are diagnosed across the globe annually, resulting in an estimated 700,000+ deaths. One major risk factor associated with development of colorectal cancer is the presence of chronic inflammation in the large intestine, also known as colitis. Inflammation is a complex immune response against harmful stimuli, characterized by symptoms including heat, redness, swelling and pain. One important molecular mediator of this process is interleukin 17 (IL-17), a pro-inflammatory …

Il-17 Drives Copper Uptake And Activation Of Growth Pathways In Colorectal Cancer Cells In A Steap4-Dependent Manner, Evan Martin

ETD Archive

Colorectal cancer is a disease characterized by abnormal, invasive cell growth beginning in the colon or rectum. The third most common type of cancer worldwide, approximately one million new cases of the disease are diagnosed across the globe annually, resulting in an estimated 700,000+ deaths. One major risk factor associated with development of colorectal cancer is the presence of chronic inflammation in the large intestine, also known as colitis. Inflammation is a complex immune response against harmful stimuli, characterized by symptoms including heat, redness, swelling and pain. One important molecular mediator of this process is interleukin 17 (IL-17), a pro-inflammatory …

Structural Characterization Of A Human/Porcine Chimeric Fviii Construct And An Improved Human Factor Viii Model And Progress Towards Determination Of The Fviii C1 Domain In Complex With Inhibitory Antibodies, Ian Smith

WWU Graduate School Collection

Blood coagulation factor VIII (FVIII) is a non-enzymatic protein cofactor, which plays a crucial role in the formation of a stable blood clot. Absence or deficiency of FVIII results in the blood disorder hemophilia A, with symptoms including internal hemorrhaging and the inability to stop bleeding from open wounds. Treatment of hemophilia A relies on replacement of FVIII with blood, plasma, or protein concentrate infusions. Unfortunately, approximately 30% of patients receiving replacement FVIII generate pathologic anti-FVIII inhibitory antibodies, which both reduce the effectiveness of the FVIII therapeutic and increase the severity of hemophilia A symptoms.

This thesis reports the determination …

Using Nucleotide Analogs As Biochemical Probes To Evaluate The Mechanisms Involved In Translesion Replication By A High Fidelity Dna Polymerase, Anvesh Dasari

ETD Archive

Translesion DNA synthesis (TLS) allows DNA polymerases to incorporate nucleotides opposite and beyond damaged DNA. This activity is an important risk factor for the initiation and progression of genetic diseases including cancer. My study evaluates the ability of a high-fidelity DNA polymerase to perform TLS with 8-oxo-guanine, a pro-mutagenic DNA lesion formed by reactive oxygen species. Using modified purine and non-natural indole analogs as biochemical probes, I have evaluated the influence of desolvation, hydrogen bonding interactions, and shape complementarity towards nucleotide binding and incorporation opposite the miscoding lesion 8-oxo-guanine by the high fidelity gp43exo- DNA polymerase. In Chapter II, I …

Expression And Purification Of Atg11: A Protein Involved In The Organization Of Autophagy, Carson Zois

Senior Honors Theses and Projects

Autophagy is a physiological process for the degradation of internal cell components to promote cell survival. This process allows the cell to maintain homeostasis, recycle damaged cellular components and helps protect against disease. Autophagy can be selective or non-selective. Non-selective, or bulk, autophagy occurs during times of cellular starvation and results in non-specific degradation of cytoplasmic materials. Selective autophagy targets specific components within the cytoplasm for degradation, and is always active. In both forms of autophagy, components are degraded when a double-membraned structure, called the autophagosome, encapsulates the target cargo and is then recruited to the lysosome in mammalian cells, …

The Rational Investigation Of Anti-Cancer Peptide Specificity Using The Knob-Socket Model, Shivarni Patel

University of the Pacific Theses and Dissertations

Cancer has been a pervasive and deadly problem for many years. No treatments have been developed that effectively destroy cancer cells while also keeping healthy cells safe. In this work, the knob-socket construct is used to analyze two systems involved in cancer pathways, the PDZ domain and the Bcl-BH3 complex. Application of the knob-socket model in mapping the packing surface topology (PST) allows a direct analysis of the residue groups important for peptide specificity and affinity in both of these systems. PDZ domains are regulatory proteins that bind the C-terminus of peptides involved in the signaling pathway of cancer progression. …

The Function Of Renalase, Brett Allen Beaupre

Theses and Dissertations

Renalase was originally reported to be an enzyme secreted into the blood by the kidney to lower blood pressure and slow heart rate. Despite multiple reports claiming to confirm this activity in vivo there has been considerable discord in regards to the reaction catalyzed by renalase. The structural topology of renalase resembles that of known flavoprotein oxidases, monooxygenases and demethylases, but the conserved active site residues are unique to renalase. It has been reported that the catalytic function of renalase is to oxidize circulating catecholamines, however in vitro studies have failed to demonstrate a catalytic activity in the presence of …

Profiling Cell Surface Sialylation And Desialylation Dynamics Of Immune Cells, Dan Wang

ETD Archive

Sialic acids (SAs) are a diverse family of naturally occurring 2-keto-3-deoxy-nononic acids that are involved in a wide range of biological processes, including early fetal development, cellular recognition, and utilization by microbes. While it is clear that cell surface SAs are highly involved in the immune system, the sialylation status of individual immune cells and functions are still unknown. In this study, I combined the newly developed LC-MS/MS methods with flow cytometry and confocal microscopy to systematically study the sialylation and desialylation dynamics of macrophages at different conditions.

First, I developed an accurate LC-MS/MS method to quantify free SA in …

Study Of Electron Transfer Through The Reductase Domain Of Neuronal Nitric Oxide Synthase And Development Of Bacterial Nitric Oxide Synthase Inhibitors, Yue Dai

ETD Archive

Crystal structure of neuronal Nitric Oxide Synthase reductase (nNOSr) implies that large-scale domain motion is essential for electron transfer. However, the details are not well understood. To address this, we generated a functioning “Cys-lite” version of nNOSr and then replaced the nNOSr Glu816 and Arg1229 residues with Cys in the FMN and FAD domains (CL5SS) in order to allow cross-domain disulfide bond formation under pH 9 or to cross-linking using bis-maleimides. Cross-linked CL5SS exhibited a =95% decrease in cytochrome c reductase activity and reduction of the disulfide bond restored the activities. The results demonstrate that a conformational equilibrium involving FMN …

Investigating The Structure Of The Papain-Inhibitor Complex Using Spr And Nmr, Margaret Sara Thomasson

LSU Doctoral Dissertations

Cysteine proteases (CPs) are enzymes with a nucleophilic thiol in their active sites. Inhibitors of cysteine proteases (ICPs) occur naturally in bacterial pathogens and some protozoa. In parasites, ICPs are often virulence factors, contributing to the formation and survival of amastigotes within host cells. These amastigotes have higher CP activity, therefore making both ICPs and CPs potential drug targets. Despite great genetic variability, ICPs contain highly conserved structural features, including a series of defined loops that play a significant role in binding CPs. Papain, a CP from Carica papaya, complexes with ICP from Leishmania mexicana. Although the individual 3-D structures …

First Steps Of Putting Research Into Practice: Utilizing Concept Inventories To Identify Biochemistry Misconceptions And The Development Of A Guided Inquiry Activity To Correct The Identified Misconceptions, Ellen Humphreys

Master of Science in Chemical Sciences Theses

It is known that students leave science classes with an incomplete or incorrect understanding of some of the concepts covered in the courses. Identification of these misconceptions is difficult, as it usually involves conducting an hour long one-on-one interview with a student. Concept inventories were developed as a way for professors to identify misconceptions in their classroom in an efficient manner. However, there is no collection of biology, chemistry and biochemistry concept inventories and there has been no analysis of the quality of these inventories. One goal of the research was to collect these inventories and do the much needed …

Purple Urine Bag Syndrome: Regulation Of Pyrimidine Enzymes, Ashley N. Broadway

Honors Theses

Purple Urine Bag Syndrome, PUBS, is a unique condition that shows an interesting discoloration of the urine from women who have a chronic catheter. This study focused on the regulation of enzyme activities to determine the effects that may create an environment where PUBS is seen in patients with catheters. In this study, the reductive pyrimidine pathway was used to read out the bacteria Pseudomonas aeruginosa ATCC 15692. This microorganism expresses a pathway that synthesizes indoxyl from tryptophan. The bacteria strains were grown using various carbon and nitrogen sources. Concentrations of acetonitrile at 2 mM and 10 mM, concentrations of …