Biochemistry, Biophysics, and Structural Biology Commons^™

Modeling The Adaptive Immune Response To Mutation-Generated Antigens, Rory J. Geyer

University Scholar Projects

Somatic mutations may drive tumorigenesis or lead to new, immunogenic epitopes (neoantigens). The immune system is thought to represses neoplastic growths through the recognition of neoantigens presented only by tumor cells. To study mutations as well as the immune response to mutation-generated antigens, we have created a conditional knockin mouse line with a gene encoding, 5’ to 3’, yellow fluorescent protein (YFP), ovalbumin (which is processed to the immunologically recognizable peptide, SIINFEKL), and cyan fluorescent protein (CFP), or, YFP-ovalbumin-CFP. A frame shift mutation has been created at the 5’ end of the ovalbumin gene, hence YFP should always be expressed, …

Molecular Mechanisms For Regulating The Assembly Of The Mixed Lineage Leukemia-1 (Mll1) Core Complex, Michael Hu

Renée Crown University Honors Thesis Projects - All

Epigenetics is the study of heritable traits that are not caused by alterations to DNA. An example of such a mechanism is histone methylation, specifically histone H3 lysine 4 (H3K4). Histones are proteins that wrap and package DNA into nucleosomes. Modifications to a histone’s chemistry can up or down regulate specific gene expression programs through changes in nucleosome positioning. Human mixed lineage leukemia protein-1 (MLL1) is a member of the SET1 family of H3K4 methyltransferases. Previous studies in the Cosgrove lab have shown that an interaction between two protein sub-complexes, MLL1 and WRAD (WDR5, RbBP5, Ash2L, DPY30), leads to dimethylation. …

The Purification And Characterization Of The Drosophila Melanogaster Trithorax Protein And Its Implications In The Studies Of The Set Domain Family Of Proteins, Kelsey Monteith

Renée Crown University Honors Thesis Projects - All

Methylation at histone H3 lysine 4 (H3K4) is a post-translational modification often associated with transcriptional regulation through altering the structural state of chromatin. The human mixed lineage leukemia protein-1 protein (MLL1) is a known histone methyltransferase that catalyzes the transfer of methyl groups to H3K4. MLL1 works in a core complex with other essential components, proteins WDR5, RbBP5, Ash2L, DPY-30 (WRAD), which is required for H3K4 dimethylation. Trithorax (TRX) protein is the Drosophila melanogaster ortholog to human MLL1, and although structurally similar is unable to perform dimethylation when in complex with the human components. The goal of this study is …

Determination Of Pore Size Distribution In Capillary-Channeled Polymer (C-Cp) Fiber Stationary Phases By Inverse Size-Exclusion Chromatography (Isec) And The Study Of The Role Of Interstitial Fraction On C-Cp Fibers On Protein Binding Capacity, Zhengxin Wang

All Theses

ABSTRACT High performance liquid chromatography (HPLC), first used in the 1960's, is a rapidly evolving analytical technique, widely employed for identification, separation, and purification in biotechnology and pharmaceutical industries. The development of the stationary phases has played an important role in improving this technique. Each stationary phase will have its own disadvantages. Polysaccharide-based stationary phases such as cross-linked dextran cannot tolerate high pressures and linear velocities; silica stationary phases are rigid enough but slow mass transfer in the pores on the surface causes another problem; with the introduction of non-porous and small bead packing materials, the low surface area and …

The Role Of Stress-Activated Protein Kinases In Parasites, Tamara Kreiss

Theses, Dissertations and Culminating Projects

Lymphatic filariasis and leishmaniasis are neglected tropical diseases that are caused by nematode and protozoal parasites. These diseases cause disfiguration, leaving their host socially marked, and in some cases cause more severe disease that can lead to death. These infections, which tend to persist for long periods of time, also lead to bacterial and fungal co-infections, which further exacerbate the disease. Currently there are insufficient treatment options. Current therapies are often too expensive, have toxicity associated with them and are subjected to growing resistance amongst parasite populations. In this thesis I investigated two potential drug targets. The first, is the …

Synthesis And Characterization Of Co(Ii) And Cu(Ii) Pyrophosphate Coordination Complexes For Treatment Of Pathogenic Agents., Leann Jade Miles

Renée Crown University Honors Thesis Projects - All

Candidiasis, the yeast infection of the mouth, genitals, and blood can be fatal in individuals who are immunocompromised. In humans, Candidiasis is most commonly caused by the yeast, Candida albicans (C. albicans), but can also be caused by other Candida species .^[1]

C. albicans demonstrates marked resistance towards Cu(II) but more sensitivity towards Co(II) compounds, especially in mutants that lack the CaCRP1 and CaCUP1 gene that encodes for the copper-transporting P-type ATPase and Metallothionein proteins, respectively.^[2] Here, I will describe the effects of Co(II) and Cu(II) metal based complexes that incorporate a bridging pyrophosphate …

Microgenomic Approaches To Identify Clinically Relevant Gene Signatures That Discriminate Histologic Types Of Breast Carcinomas., Sean Thomas Butterbaugh

College of Arts & Sciences Senior Theses

Background: Breast cancer presents itself in a variety of histologic types, and the two most common types are invasive ductal carcinoma (IDC) and invasive lobular carcinoma (ILC). Based on comparative genomic hybridization (CGH) analyses, ILC is more closely related to low grade IDC than it is to intermediate and high grade IDC. Results from the BIG 1-98 trial demonstrate that post-menopausal women who are affected with estrogen receptor positive (ER+) ILC or luminal B (high grade) IDC experience a greater magnitude of benefit when they are treated with the aromatase inhibitor (AI) letrozole compared to treatment with the antiestrogen tamoxifen. …

Porphyrin Cross-Linkers For Generating Soluble Molecularly Imprinted Polymers From Polyethyleneimine, Mohammed R. Elshaer

Seton Hall University Dissertations and Theses (ETDs)

Molecular recognition is vital to many biochemical processes and is at the heart of promising bio-medically related technologies. Molecular imprinting has a long-standing history as a successful method for mimicking the molecular recognition phenomena exhibited by nature, whereby artificial receptors are prepared for a given target molecule based on synthetic polymers. The molecularly imprinted polymer (MIP) contains a three dimensional network with a memorized cavity specific to the shape and functionality of the templated target molecule. The utility of traditional MIPs has been limited due to an inherent lack of solubility. We have worked toward developing a system that allows …

Expression Of Human Neutrophil Cathepsin G In Pichia Pastoris, Evan T. Perry

Undergraduate Honors Theses

Cathepsin G (CatG), a serine protease found in the azurophil granules of neutrophils, participates in killing engulfed microorganisms. CatG is a poorly understood enzyme, in part because it can only be obtained as mature enzyme purified from human blood, and because it seems to have dual specificity for chymotrypsin-like and trypsin-like substrates. Therefore, yeast Pichia pastoris was used to express immature recombinant human CatG to provide a source of the enzyme free of biohazards and to allow study of its dual specificity and C-terminal processing. To avoid potential cleavage by a yeast kexin protease, the amino sequence was modified without …

Control Of Cellular Motility By Neuropilin-Mediated Physical Interactions, Xiaobo Li, Matthew W. Parker, Craig W. Vander Kooi

Molecular and Cellular Biochemistry Faculty Publications

The neuropilin (Nrp) family consists of multifunctional cell surface receptors with critical roles in a number of different cell and tissue types. A core aspect of Nrp function is in ligand-dependent cellular migration, where it controls the multistep process of cellular motility through integration of ligand binding and receptor signaling. At a molecular level, the role of Nrp in migration is intimately connected to the control of adhesive interactions and cytoskeletal reorganization. Here, we review the physiological role of Nrp in cellular adhesion and motility in the cardiovascular and nervous systems. We also discuss the emerging pathological role of Nrp …

Propagation Of Oligomeric Α-Synuclein And Amyloid-Β: Implications For Parkinson's And Alzheimer's Diseases, Matthew Stephen Planchard

Master's Theses

The aggregation of amyloidogenic proteins is a critical event in the pathology of a variety of neurodegenerative diseases, including Alzheimer’s disease (AD) and Parkinson’s disease (PD). The proteins α-synuclein (αS) and amyloid-β (Aβ) are involved in the formation of amyloid lesions observed in PD and AD, respectively. Both PD and AD exhibit a significant amount of co-pathology in clinical settings, and the αS and Aβ proteins have been shown to interact in vitro. Recent experimental consensus has shown oligomeric species to be significant, if not primary, sources of toxicity in these diseases. …

The Use Of A Small Molecule To Improve The Thermostability Of Dna Junctions, Arik Shams

Honors Theses

The short-term goal of this research project is to employ small molecules as a means to stabilize four-way DNA junctions (4WJs) composed of natural DNA and chimeric nucleic acids. The long-term goal of the project is utilizing the 4WJs as extracellular therapeutic inhibitors of DNA binding proteins [i.e. Histones and High Mobility Group Protein B (HMGB1b)]. A number of studies have shown that classical intracellular DNA-binding proteins have a variety of deleterious side-effects when present in the extracellular milieu. In order to develop a successful 4WJ therapeutic, we are focused on using modified nucleic acids to enhance the stability of …

Cloning, Expression And Interaction Studies Of The Potential Rubisco Activase Cbbq, Salma A. Dawoud

Honors Theses

Carboxysomes are polyhedral microcompartments found in all cyanobacteria and in many chemoautotrophs. Within their shell they contain the enzyme ribulose-1, 5-bisphosphate carboxylase/oxygenase (RubisCO), which fixes CO₂. Downstream of the carboxysome operon there is another gene cluster containing several genes that may enhance carboxysome function. Two of these genes, cbbQ and cbbO, encode potential RubisCO activases. Using recombinant CbbQ and CbbO protein, and RubisCO isolated from carboxysomes the interaction between these proteins was studied. The CbbO and CbbQ proteins were both His tagged, allowing them to be purified with Ni²⁺-NTA column chromatography. Each of these tagged …

The Anti-Obesogenic Effects Of Nitric Oxide., Brian Edward Sansbury 1985-

Electronic Theses and Dissertations

Obesity is a strong risk factor for developing type 2 diabetes and cardiovascular disease and has quickly reached epidemic proportions with few tangible and safe treatment options. While it is generally accepted that the primary cause of obesity is energy imbalance, i.e., more calories are consumed than are utilized, understanding how caloric balance is regulated has proven a challenge. Molecular processes and pathways that directly regulate energy metabolism represent promising targets for therapy. In particular, nitric oxide (NO) is emerging as a central regulator of energy metabolism and body composition. NO bioavailability is decreased in animal models of obesity and …

Redox Active Motifs In Selenoproteins, Fei Li, Yuliya Pepelyayeva, Elias S. J. Arner, Craig A. Bayse, Sharon Rozovsky

Chemistry & Biochemistry Faculty Publications

Selenoproteins use the rare amino acid selenocysteine (Sec) to act as the first line of defense against oxidants, which are linked to aging, cancer, and neurodegenerative diseases. Many selenoproteins are oxidoreductases in which the reactive Sec is connected to a neighboring Cys and able to form a ring. These Sec-containing redox motifs govern much of the reactivity of selenoproteins. To study their fundamental properties, we have used Se-77 NMR spectroscopy in concert with theoretical calculations to determine the conformational preferences and mobility of representative motifs. This use of Se-77 as a probe enables the direct recording of the properties of …

Tht And Capillary Electrophoresis To Monitor The Effects Of Solutions Conditions On Amylin Aggregation, Michael May

Graduate Theses and Dissertations

Amylin (hIAPP) aggregates have been found in 90% of patients with type II diabetes at autopsy, and are suspected to play a role in the death of islet &beta-cells¹. However, this aggregation process is not well understood. Here, we explore methods that utilize capillary electrophoresis (CE) as a means to better understand amylin's aggregation process.

We examined the effects of solutions conditions: agitation, pH, salt, and temperature on amylin aggregation using Thioflavin T, dot blots, and capillary electrophoresis. Thiofalvin T was used to predict the lag time to &beta-sheet formation. Our results indicated all variables with the exception …

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

Department of Biochemistry Faculty Publications

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

Numerical Assessment Of Sequence Conservation In Flu-Virus Hemagglutinin, Scott S. Norton

Honors Scholar Theses

The flu virus was investigated to find a common recognition domain to which an antibody against human-infected viruses can bind. If such a target site is structurally and electrostaticly conserved or invariant, only a single antibody would be required to attack the virus in all cases. The sequence of one of the viral surface proteins contains 24 amino acids that do not vary through mutation. However, these amino acids are neither contiguous in sequence or in space, and the ones that are associated with each other are not readily accessible to an antibody. They do provide a first impression of …

Characterization Of Udp-Arabinopyranose Mutase Genes In The Arabidopsis Cell Wall Mutant Mur5, Christopher A. Hart

Honors Scholar Theses

The genome of Arabidopsis thaliana contains several coding regions for UDP-arabinopyranose mutases (UAMs) that are also known as reversibly glycosylated polypeptides (RGPs). The mur5 cell wall mutant of Arabidopsis shows a 30% decrease in cell wall arabinose content, and a missense mutation in the Reversibly Glycosylated Polypeptide 2 gene was recently proposed to cause this mutant phenotype. Through a traditional complementation analysis, mur5 and a T-DNA insertion mutant in the RGP2 gene were shown not to complement each other, indicating that the two genes are mutant alleles of the same locus. The mur5 SNP located in RGP2 caused a more …

Modeling The Adaptive Immune Response To Mutation-Generated Antigens, Rory J. Geyer

Honors Scholar Theses

Somatic mutations may drive tumorigenesis or lead to new, immunogenic epitopes (neoantigens). The immune system is thought to represses neoplastic growths through the recognition of neoantigens presented only by tumor cells. To study mutations as well as the immune response to mutation-generated antigens, we have created a conditional knockin mouse line with a gene encoding, 5’ to 3’, yellow fluorescent protein (YFP), ovalbumin (which is processed to the immunologically recognizable peptide, SIINFEKL), and cyan fluorescent protein (CFP), or, YFP-ovalbumin-CFP. A frame shift mutation has been created at the 5’ end of the ovalbumin gene, hence YFP should always be expressed, …