Biochemistry, Biophysics, and Structural Biology Commons^™

Prep-Hplc Method Development To Isolate Potentially-Carcinogenic, Arginine-Based Heterocyclic Amines, Victoria S. Kim

Honors Theses

Heterocyclic amines (HCAs) are carcinogenic molecules generated from the reaction of creatin(in)e and amino acids at high cooking temperatures in meat. Previous research has shown that replacing creatine with arginine leads to new, uncharacterized HCA molecules. This research entails implementing and optimizing the preparative HPLC analysis of extracts from multiple burnings of arginine with phenylalanine. Isolated fractions from the prep-HPLC analysis were further analyzed using the Ames test to identify mutagenic compounds. Prep-HPLC provides quality control to the burning process along with isolation of larger quantities of materials for further characterization and molecular structure identification.

Cytochrome B5 Protects Photoreceptors From Light Stress-Induced Lipid Peroxidation And Retinal Degeneration, Xinping Chen, Hana Hall, Jeffrey P. Simpson, Walter D. Leon-Salas, Donald F. Ready, Vikki M. Weake

Department of Biochemistry Faculty Publications

Lipid peroxides are generated by oxidative stress in cells, and contribute to ageing and neurodegenerative disease. The eye is at special risk for lipid peroxidation because photoreceptors possess amplified sensory membranes rich in peroxidation-susceptible polyunsaturated fatty acids. Light-induced lipid peroxidation in the retina contributes to retinal degeneration, and lipid peroxidation has been implicated in the progression of age-associated ocular diseases such as age-related macular degeneration (AMD). Here, we show that exposing Drosophila melanogaster to strong blue light induces oxidative stress including lipid peroxidation that results in retinal degeneration. Surprisingly, very young flies are resilient to this acute light stress, suggesting …

Acyl-Homoserine Lactone Based Modulators For Rhli, A Quorum Sensing Signal Synthase In Pseudomonas Aeruginosa, Daniel D. Shin

Boise State University Theses and Dissertations

Gram-negative bacteria use N-acyl-homoserine lactone (AHL) autoinducer based signal system, known as quorum sensing (QS), to modulate the gene expression for such traits as biofilm formation, toxin production, and antibiotic resistance. Therefore, there is great potential in pursuing quorum sensing inhibition (QSI) as a means of achieving antivirulence. Pseudomonas aeruginosa, an opportunistic pathogen commonly found in healthcare-related infections, use two LuxI/R type systems to regulate AHL-based quorum sensing: LasI/R and RhlI/R. LasI (initiator protein/signal synthase) and LasR (receptor) use 3-oxododecanoyl-L-homoserine lactone signal molecule while RhlI and RhlR use butanoyl-L-homoserine lactone autoinducer. Thus far, most of the studies have …

Type Ix Secretion System: Characterization Of An Effector Protein And An Insight Into The Role Of C-Terminal Domain Dimeration In Outer Membrane Translocation., Lahari Koneru

Electronic Theses and Dissertations

Porphyromonas gingivalis and Tannerella forsythia are two of the primary pathogens that are associated in the etiology and progression of chronic periodontitis. In T. forsythia, KLIKK proteases are the recently identified group of proteolytic enzymes that are secreted through Type IX secretion system (T9SS). Among, these KLIKK proteases a synergistic relationship was observed between karilysin and mirolysin in invading the host complement system for the survival of the bacteria. Since, karilysin has been already characterized, in this study we propose to study about mirolysin through structural, biochemical and biological characterization. The obtained results from the experiments has shown the …

Molecular Geometries And Other Properties Of H2o⋯Agi And H3n⋯Agi As Characterised By Rotational Spectroscopy And Ab Initio Calculations, Chris Medcraft, Eva Gougoula, Dror M. Bittner, John C. Mullaney, Susana Blanco, David P. Tew, Nicholas R. Walker, Anthony C. Legon

Chemistry & Biochemistry Faculty Publications

The rotational spectra of H₃N⋯AgI and H₂O⋯AgI have been recorded between 6.5 and 18.5 GHz by chirped-pulse Fourier-transform microwave spectroscopy. The complexes were generated through laser vaporisation of a solid target of silver or silver iodide in the presence of an argon gas pulse containing a low concentration of the Lewis base. The gaseous sample subsequently undergoes supersonic expansion which results in cooling of rotational and vibrational motions such that weakly bound complexes can form within the expanding gas jet. Spectroscopic parameters have been determined for eight isotopologues of H₃N⋯AgI and six isotopologues of …

Insights Into The Therapeutic Potential Of Salt Inducible Kinase 1: A Novel Mechanism Of Metabolic Control, Randi Fitzgibbon

Dissertations and Theses (Open Access)

Salt inducible kinase 1 (SIK1) has been considered a stress-inducible kinase since it was first cloned in 1999. Continued efforts since this time have been dedicated to characterizing the structure and function of SIK1. Such research has laid the ground work for our understanding of SIK1 action and regulation in tissue and stimuli dependent manners. The fundamental findings of this dissertation continue in this tradition and include investigations of SIK1 regulatory mechanisms in skeletal muscle cells, the cellular and physiological effects of SIK1 loss of function in vitro and in vivo, and intracellular metabolic and mitochondrial regulation by this …

Role Of Cannabinoid Receptor Type 2 (Cb2) In Late Stage Atherosclerosis, Makenzie Fulmer

Electronic Theses and Dissertations

Atherosclerosis is a chronic inflammatory disorder of medium and large vessels. Immune signaling and dyslipidemia are two of several processes which influence lesion development in atherosclerosis. Cannabinoids, such as those found in marijuana, exert their effects through two cannabinoid receptors, CB1 and CB2. Recent studies using CB2 knockout mice and CB2-selective ligands have shed light on a protective role of CB2 in early stages of atherosclerosis. However, the role of CB2 in advanced stages of atherosclerosis remains unclear. To determine if CB2 plays a role in advanced atherosclerotic lesion composition and progression, we investigated the effects of systemic CB2 gene …

A Potential Solution To A Poopy Problem: Bile Salt Analogs As Prophylactics Against Clostridium Difficile Infection, Jacqueline Renee Phan

UNLV Theses, Dissertations, Professional Papers, and Capstones

Clostridium difficile infection (CDI) is a major cause of antibiotic-associated diarrhea. In 2011, over 500,000 patients were diagnosed with CDI in the United States and over 29,000 people died of CDI-related complications. With an average of $35,000 to treat a single case of inpatient CDI, cost burden to the healthcare system can reach up to $3.2 billion annually. As both hospital- and community-acquired CDI incidences rise due to the emergence of hypervirulent strains and CDI reoccurrences of up to 25%, standard treatments are rendered less effective and new methods of prevention are critical.

CDI is caused by bacteria called Clostridium …

Determination Of The Effects Of Different Maillard Reaction Products On The Taxonomic Composition Of The Gut Microbiota, Nesreen Hamdan Aljahdali

Graduate Theses and Dissertations

The Maillard Reaction (MR) is a non-enzymatic chemical reaction which results in linkage between the amino group of amino acids and the carbonyl group of reduced sugars. This reaction generates Maillard reaction products (MRPs) which are not present naturally in foods, and are responsible for a range of colors, odors, flavors, and other sensory properties. Conflicting reports of MRPs impacts on human health are probably due to the fact that bioconversion of these digestible molecules by the gut microbiota has been marginally taken into account. This study aimed to determine the effects of different MRPs on rodent’s gut microbiota through16S …

Harmonizing Lipidomics: Nist Interlaboratory Comparison Exercise For Lipidomics Using Srm 1950-Metabolites In Frozen Human Plasma, J Bowden, C Ulmer, C Jones, J Koelmel, L Abdullah, Houli Jiang, Michal Schwartzman, Amaury Cazenave-Gassiot, Antonio Checa, Michelle Cinel, Romain Colas, Serge Cremers, Edward Dennis, James Evans, Alexander Fauland, Jun Han, Houli Jiang, Michal Schwartzman

NYMC Faculty Publications

As the lipidomics field continues to advance, self-evaluation within the community is critical. Here, we performed an interlaboratory comparison exercise for lipidomics using Standard Reference Material (SRM) 1950-Metabolites in Frozen Human Plasma, a commercially available reference material. The interlaboratory study comprised 31 diverse laboratories, with each laboratory using a different lipidomics workflow. A total of 1,527 unique lipids were measured across all laboratories and consensus location estimates and associated uncertainties were determined for 339 of these lipids measured at the sum composition level by five or more participating laboratories. These evaluated lipids detected in SRM 1950 serve as community-wide benchmarks …

Hydrogen-Bonding-Directed Ordered Assembly Of Carboxylated Poly(3-Alkylthiophene)S, David W. Bilger, Jose A. Figueroa, Neil D. Redeker, Amrita Sarkar, Morgan Stefik, Shanjuu Zhang

Faculty Publications

Hydrogen-bonding-induced ordered assembly of poly(3-alkylthiophene)s derivatives bearing carboxylic acid groups has been investigated from diluted and concentrated solutions to solid films using ultraviolet–visible absorption spectroscopy, polarized optical microscopy, and four-point probe conductivity measurements. In dilute solutions, the polymer undergoes a spontaneous structural transition from disordered coil-like to ordered rodlike conformations, which is evidenced by time-dependent chromism. Many factors such as alkyl-chain length, types of solvents, and temperature are studied to understand the assembly behavior. Transition kinetics of the assembly process reveals a universal second-order rate law, indicating an intermolecular origin due to hydrogen bonding. When more concentrated, hydrogen bonding drives …

Investigations Into The Non-Mevalonate Isoprenoid Biosynthesis Pathway's First Two Enzymes Utilizing Hybrid Qm/Mm Techniques, Justin K. White

USF Tampa Graduate Theses and Dissertations

Molecular drug design begins with the identication of a problem to solve. This work identies the growing resistance among human pathogens to current treatments. Once the problem is identied and understood, solutions must be proposed. This one is straight forward, we need new antimicrobial drugs. More specically, we need to identify novel targets to inhibit. A large portion of antibiotics focus on disruption of macromolecular production while only a few target metabolic systems. Finally, you need to propose solutions based on the information gathered. In order to avoid existing resistance, it is important to avoid the macromolecular route and focus …

Design And Synthesis Of Cpg-Lytic Peptide Conjugate, Brachytherapy Beads And A Combinatorial Library Of Primary Amines Used As Potential Therapeutics In The Treatment Of Cancers, Josanne-Dee Woodroffe

USF Tampa Graduate Theses and Dissertations

Cancer remains one of the most feared diseases affecting the modern world. Second to heart disease, it is the largest cause of deaths, affecting one in three persons. Cancer cells are formed when normal, healthy cells become damage, losing their normal regulatory mechanism that control cell growth. There are many different types and progression of these cancer cells that determine the type of treatment a patient receives. The primary focus of this dissertation is to propose three studies of anticancer agents. In Chapter one, a CpG-lytic peptide conjugate was designed to target receptors on the cell membrane to concentrate the …

Mechanism Elucidation And Inhibitor Discovery Against Serine And Metallo-Beta-Lactamases Involved In Bacterial Antibiotic Resistance, Orville A. Pemberton

USF Tampa Graduate Theses and Dissertations

The emergence and proliferation of Gram-negative bacteria expressing β-lactamases is a significant threat to human health. β-Lactamases are enzymes that degrade the β-lactam antibiotics (e.g., penicillins, cephalosporins, monobactams, and carbapenems) that we use to treat a diverse range of bacterial infections. Specifically, β-lactamases catalyze a hydrolysis reaction where the β-lactam ring common to all β-lactam antibiotics and responsible for their antibacterial activity, is opened, leaving an inactive drug. There are two groups of β-lactamases: serine enzymes that use an active site serine residue for β-lactam hydrolysis and metalloenzymes that use either one or two zinc ions for catalysis. Serine enzymes …

Disorder Levels Of C-Myb Transactivation Domain Regulate Its Binding Affinity To The Kix Domain Of Creb Binding Protein, Anusha Poosapati

USF Tampa Graduate Theses and Dissertations

Intrinsically disordered proteins (IDPs) do not form stable tertiary structures like their ordered partners. They exist as heterogeneous ensembles that fluctuate over a time scale. Intrinsically disordered regions and proteins are found across different phyla and exert crucial biological functions. They exhibit transient secondary structures in their free state and become folded upon binding to their protein partners via a mechanism called coupled folding and binding. Some IDPs form alpha helices when bound to their protein partners. We observed a set of cancer associated IDPs where the helical binding segments of IDPs are flanked by prolines on both the sides. …

Investigating The Toxicity And Accumulation Of 2,4-Dichlorophenoxyacetic Acid (2,4-D) And Glyphosate In Eisenia Fetida, Caitlin Lazurick, Nicole Lidzbarski, Rachel Owings, Jeff Brotherton, Edna Steele

Journal of the South Carolina Academy of Science

Glyphosate, the main ingredient in Roundup^®, is the most widely used herbicide in the world. It is also used in combination with another effective herbicide, 2,4-D, in the formulation called Enlist Duo^®. The EPA approved the use of Enlist Duo^® on certain crops including those genetically modified to be resistant to both herbicides. The predicted significant increase in the use of these herbicides raised concerns from the general public because both compounds have been classified as possibly or probably carcinogenic. Since herbicides are applied directly to vegetation, the safety of organisms that come in contact …

Defining Electron Bifurcation In The Electron-Transferring Flavoprotein Family, Amaya M. Garcia Costas, Saroj Poudel, Anne-Frances Miller, Gerrit J. Schut, Rhesa N. Ledbetter, Kathryn R. Fixen, Lance C. Seefeldt, Michael W. W. Adams, Caroline S. Harwood, Eric S. Boyd, John W. Peters

Chemistry Faculty Publications

Electron bifurcation is the coupling of exergonic and endergonic redox reactions to simultaneously generate (or utilize) low- and high-potential electrons. It is the third recognized form of energy conservation in biology and was recently described for select electron-transferring flavoproteins (Etfs). Etfs are flavin-containing heterodimers best known for donating electrons derived from fatty acid and amino acid oxidation to an electron transfer respiratory chain via Etf-quinone oxidoreductase. Canonical examples contain a flavin adenine dinucleotide (FAD) that is involved in electron transfer, as well as a non-redox-active AMP. However, Etfs demonstrated to bifurcate electrons contain a second FAD in place of the …

The Future Perspective: Metabolomics In Laboratory Medicine For Inborn Errors Of Metabolism, Yana Sandlers

Chemistry Faculty Publications

Metabolomics can be described as a simultaneous and comprehensive analysis of small molecules in a biological sample. Recent technological and bioinformatics advances have facilitated large-scale metabolomic studies in many areas, including inborn errors of metabolism (IEMs). Despite significant improvements in the diagnosis and treatment of some IEMs, it is still challenging to understand how genetic variation affects disease progression and susceptibility. In addition, a search for new more personalized therapies and a growing demand for tools to monitor the long-term metabolic effects of existing therapies set the stage for metabolomics integration in preclinical and clinical studies. While targeted metabolomics approach …

Regenerative Therapy By Replacement Of Stem Cells In Females, Saara Ahmad, Rehana Rehman, Fatima Ahmad

Department of Biological & Biomedical Sciences

No abstract provided.

Association Of Vitamin D Deficiency And Vdbp Gene Polymorphism With The Risk Of Ami In A Pakistani Population, Mujtaba Mubashir, Shaheena Anwar, Asal Khan Tareen, Naseema Mehboobali, Khalida Iqbal, Mohammad Iqbal

Department of Biological & Biomedical Sciences

OBJECTIVE: To investigate the relationship of vitamin D deficiency and risk of AMI in a Pakistani population, and to find out any associationbetween vitamin D binding protein (VDBP) genotypes and risk of AMI in this population.
METHODS: In a comparative cross-sectional study, 246 patients (age: 20-70 years; 171 males and 75 females) with first AMI were enrolled with informed consent. Similarly, 345 healthy adults (230 males and 115 females) were enrolled as controls. Their fasting serum samples were analyzed for 25 (OH) vitamin D, lipids and other biomarkers using kit methods, while DNA was analyzed for VDBP genotypes using PCR-RFLP …

Mitochondrial Reactive Oxygen Species In Lipotoxic Hearts Induces Post-Translational Modifications Of Akap121, Drp1 And Opa1 That Promote Mitochondrial Fission, Kensuke Tsushima, Heiko Bugger, Adam R. Wende, Jamie Soto, Gregory A. Jenson, Austin R. Tor, Rose Mcglauflin, Helena C. Kenny, Yuan Zhang, Rhonda Souvenir, Xiao X. Hu, Crystal L. Sloan, Renata O. Pereira, Vitor A. Lira, Kenneth W. Spitzer, Terry L. Sharp, Kooresh I. Shoghi, Genevieve C. Sparagna, Eva A. Rog-Zielinska, Peter Kohl, Oleh Khalimonchuk, Jean E. Schaffer, E. Dale Abel

Department of Biochemistry: Faculty Publications

Rationale: Cardiac lipotoxicity, characterized by increased uptake, oxidation and accumulation of lipid intermediates, contributes to cardiac dysfunction in obesity and diabetes. However, mechanisms linking lipid overload and mitochondrial dysfunction are incompletely understood.

Objective: To elucidate the mechanisms for mitochondrial adaptations to lipid overload in postnatal hearts in vivo.

Methods and Results: Using a transgenic mouse model of cardiac lipotoxicity overexpressing long-chain acyl-CoA synthetase 1 in cardiomyocytes, we show that modestly increased myocardial fatty acid uptake leads to mitochondrial structural remodeling with significant reduction in minimum diameter. This is associated with increased palmitoyl-carnitine oxidation and increased reactive oxygen species (ROS) generation …

Association Of Vitamin D Binding Protein Polymorphism With Risk Of Type 2 Diabetes Mellitus In A Pakistani Urban Population: A Case Control Study, Khalida Iqbal, Najmul Islam, Iqbal Azam, Ali Asghar, Naseema Mehboobali, Mohammad Perwaiz Iqbal

Department of Biological & Biomedical Sciences

Objective: To assess if genotypes/diplotypes of vitamin D binding protein have any association with type 2 diabetes mellitus.
Methods: This case-control study was conducted from January 2013 to July 2015 at the endocrinology clinics of the Aga Khan University Hospital, Karachi, and comprised adult patients with type 2 diabetes and their age- and gender-matched healthy controls. Venous blood was obtained and assessed for serum/plasma 25 hydroxyvitamin D, parathyroid hormone, calcium, alkaline phosphatase and creatinine. Deoxyribonucleic acid was isolated and genotyping was done by polymerase chain reaction-restriction fragment length polymorphismprocedures.
Results: Of the 330 participants, there were 165(50%) …

High-Throughput Single-Molecule Telomere Characterization, Jennifer Mccaffrey, Eleanor Young, Katy Lassahn, Justin Sibert, Steven Pastor, Harold Riethman, Ming Xiao

School of Medical Diagnostics & Translational Sciences Publications

We have developed a novel method that enables global subtelomere and haplotype-resolved analysis of telomere lengths at the single-molecule level. An in vitro CRISPR/Cas9 RNA-directed nickase system directs the specific labeling of human (TTAGGG) n DNA tracts in genomes that have also been barcoded using a separate nickase enzyme that recognizes a 7bp motif genome-wide. High-throughput imaging and analysis of large DNA single molecules from genomes labeled in this fashion using a nanochannel array system permits mapping through subtelomere repeat element (SRE) regions to unique chromosomal DNA while simultaneously measuring the (TTAGGG) n tract length at the end of each …

Using Competition Assays To Quantitatively Model Cooperative Binding By Transcription Factors And Other Ligands., Jacob Peacock, James B Jaynes

Department of Biochemistry and Molecular Biology Faculty Papers

BACKGROUND: The affinities of DNA binding proteins for target sites can be used to model the regulation of gene expression. These proteins can bind to DNA cooperatively, strongly impacting their affinity and specificity. However, current methods for measuring cooperativity do not provide the means to accurately predict binding behavior over a wide range of concentrations.

METHODS: We use standard computational and mathematical methods, and develop novel methods as described in Results.

RESULTS: We explore some complexities of cooperative binding, and develop an improved method for relating in vitro measurements to in vivo function, based on ternary complex formation. We derive …

Redox-Active Nanomaterials For Nanomedicine Applications, C. M. Sims, S. K. Hanna, D. A. Heller, C. P. Horoszko, M. E. Johnson, A. R. Montoro Bustos, V. Reipa, Kathryn R. Riley, B. C. Nelson

Chemistry & Biochemistry Faculty Works

Nanomedicine utilizes the remarkable properties of nanomaterials for the diagnosis, treatment, and prevention of disease. Many of these nanomaterials have been shown to have robust antioxidative properties, potentially functioning as strong scavengers of reactive oxygen species. Conversely, several nanomaterials have also been shown to promote the generation of reactive oxygen species, which may precipitate the onset of oxidative stress, a state that is thought to contribute to the development of a variety of adverse conditions. As such, the impacts of nanomaterials on biological entities are often associated with and influenced by their specific redox properties. In this review, we overview …

Kinetic Characterization Of Listeria Monocytogenes 2-C- Methyl-D-Erythritol 4-Phosphate Cytidylyltransferase ( Cms ) Enzymes Using High Performance Liquid Chromatography ( Hplc ), Mark Oblazny

Theses and Dissertations

Infectious diseases, with increasing prevalence of antibiotic resistant bacteria, coupled with the declining rate in discovery of antimicrobial agents, impose one of the most significant threats to human health. Here we identify 2-C-methyl-D-erythritol 4-phosphate cytidylyltransferase (CMS) as a valid target for antibiotic development which is an enzyme in pathogenic organisms that leads to the biosynthesis of isoprene precursor molecules. Isoprene molecules are one of nature’s most common building blocks that are vital to many biological metabolic processes and are synthesized via the mevalonic acid dependent (MVA), or methylerythritol phosphate (MEP) pathway. Vertebrates utilize the MVA pathway, while many pathogenic bacteria …

The Biochemical Assessment Of Two Secreted Acid Phosphatases From Leishmania Tarentolae, Their Response To Electric Fields, Glycosidase Incubation, And / Or Vanadium, Benjamin M. Dorsey

Theses and Dissertations

Leishmaniasis, as defined by the Center for Disease Control and Prevention, is a neglected tropical disease with 1.6 million new cases reported each year. However, there is yet to be safe, effective, and affordable treatments provided to those affected by this disease1. Still underappreciated as a potential pharmaceutical targets, especially for cutaneous leishmaniasis infections, are the two isozymes of secreted acid phosphatase (SAP); secreted acid phosphatase 1 (SAP1) and secreted acid phosphatase 2 (SAP2). These enzymes are involved in the survival of the parasite in the sand fly vector, and the prevention of host macrophages from forming parasitophorous vacuole and …

Prebiotic Rna Network Formation: A Taxonomy Of Molecular Cooperation, Cole Mathis, Sanjay N. Ramprasad, Sara Imari Walker, Niles Lehman

Chemistry Faculty Publications and Presentations

Cooperation is essential for evolution of biological complexity. Recent work has shown game theoretic arguments, commonly used to model biological cooperation, can also illuminate the dynamics of chemical systems. Here we investigate the types of cooperation possible in a real RNA system based on the Azoarcusribozyme, by constructing a taxonomy of possible cooperative groups. We construct a computational model of this system to investigate the features of the real system promoting cooperation. We find triplet interactions among genotypes are intrinsically biased towards cooperation due to the particular distribution of catalytic rate constants measured empirically in the real system. For other …

Functional Characteristics And Genomic Sequencing Of Antarctic Environmental Isolates, Amanda Kemper, Christina Aigner, Michelle Tigges, Sara Anderson

Annual Symposium on Biomathematics and Ecology Education and Research

No abstract provided.

2-D Electrophoresis Modeling Of Multienzyme Cutting Of Polypeptides, Howard Mayes

Theses

2-Dimensional Electrophoresis is one of the tools in the identification of proteins by molecular weight and pH. The display of molecular weight allows the researcher to quickly identify whether a specific protein or peptide string is in the sample. The pH measurement allows even better resolution between different species in the sample. The MultiEnzyme ElectroPhoresis (MEEP) program tries to model that by providing a graph that displays separated protein strings by both molecular weight and pH. The ability to cleave the protein with 43 different enzyme variations allows the researcher to analyze appropriate enzymes to isolate a protein subsequence before …