Biochemistry, Biophysics, and Structural Biology Commons^™

Substrate Channeling In Proline Metabolism, Benjamin W. Arentson, Nikhilesh Sanyal, Donald F. Becker

Department of Biochemistry: Faculty Publications

Proline metabolism is an important pathway that has relevance in several cellular functions such as redox balance, apoptosis, and cell survival. Results from different groups have indicated that substrate channeling of proline metabolic intermediates may be a critical mechanism. One intermediate is pyrroline-5-carboxylate (P5C), which upon hydrolysis opens to glutamic semialdehyde (GSA). Recent structural and kinetic evidence indicate substrate channeling of P5C/ GSA occurs in the proline catabolic pathway between the proline dehydrogenase and P5C dehydrogenase active sites of bifunctional proline utilization A (PutA). Substrate channeling in PutA is proposed to facilitate the hydrolysis of P5C to GSA which is …

Expansion Dating: Calibrating Molecular Clocks In Marine Species From Expansions Onto The Sunda Shelf Following The Last Glacial Maximum, Eric D. Crandall, Elizabeth J. Sbrocco, Timery S. Deboer, Paul H. Barber, Kent E. Carpenter

Biological Sciences Faculty Publications

The rate of change in DNA is an important parameter for understanding molecular evolution and hence for inferences drawn from studies of phylogeography and phylogenetics. Most rate calibrations for mitochondrial coding regions in marine species have been made from divergence dating for fossils and vicariant events older than 1-2 My and are typically 0.5-2% per lineage per million years. Recently, calibrations made with ancient DNA (aDNA) from younger dates have yielded faster rates, suggesting that estimates of the molecular rate of change depend on the time of calibration, decaying from the instantaneous mutation rate to the phylogenetic substitution rate. aDNA …

Non-Canonical Roles For Stat1 And Stat2 In Mitochondrial Biogenesis And Fasting Homeostasis., Jennifer Sisler

Theses and Dissertations

The signal transducer and activator of transcription 1 (STAT1) and 2 (STAT2) are primarily activated by interferons and play a central role in orchestrating responses to a variety of pathogens by activating the transcription of nuclear encoded genes that mediate antiviral, antigrowth and immune surveillance responses. In addition to their nuclear effects, we report that STAT1 and STAT2 inhibit the expression of mitochondrial encoded mRNAs by activating type I interferons (IFNβ) under basal conditions. STAT1-/- livers also exhibit elevated levels of nuclear encoded components of the electron transport chain (ETC). Treatment of wild-type mice with IFNβ inhibits both mitochondrial and …

Identifying Sortase A Variants With Higher Catalytic Effeciency, Muna Suliman

Theses and Dissertations

In the past two decades, the field of protein engineering has evolved rapidly to include new genetic and chemical techniques to alter protein function. Protein engineering seeks to improve enzyme properties through powerful methods that specifically incorporate novel or improved function in proteins. One such method is protein ligation, which is used to selectively link synthetic and recombinant polypeptides. Due to the limitations of current protein labeling techniques, simple site-specific modification methods remain in high demand. Use of enzyme-based labeling has been the focus of various studies because of its substrate specificity. Sortase-mediated transpeptidation is one approach that has been …

Role Of Lysophosphatidic Acid In Regulation Of Cancer Cell Metabolism, Abir Mukherjee

Theses and Dissertations

The simplest phospholipid, lysophosphatidic acid (LPA), is a heat stable component of serum known for its proliferative and migratory activities in cancer cells. Strong evidence suggests that LPA production and expression of its receptors are dysregulated in multiple human malignancies. The mechanism behind LPA-mediated tumor cell growth and oncogenesis remains poorly understood. In this thesis project I used ovarian and other cancer cells as a model system to examine the hypothesis that LPA present in the tumor microenvironment is a pathophysiological determinant of hyperactive de novo lipogenesis and aerobic glycolysis, two hallmarks of cancer cells. We demonstrated that LPA induced …

Enantioselective Demethylation: The Key To The Nornicotine Enantiomeric Composition In Tobacco Leaf, Bin Cai

Theses and Dissertations--Plant and Soil Sciences

Nicotine and nornicotine are the two main alkaloids that accumulate in Nicotiana tabacum L. (tobacco), and nornicotine is the N-demethylation metabolite of nicotine. Nicotine is synthesized in the root, and probably primarily in the root tip. Both nicotine and nornicotine exist as two isomers that differ from each other by the orientation of H atom at the C-2' position on the pyrrolidine ring. (S)-nicotine is the dominant form in tobacco leaf and the enantiomer fraction of nicotine (EF_nic), the fraction of (R)-enantiomer over the total nicotine, is approximately 0.002. Despite considerable efforts to elucidate nicotine and nornicotine …

The Role Of R7 Regulators Of G Protein Signaling In The Retina, Hoon Shim

Theses and Dissertations

The R7 regulators of G protein signaling (R7 RGS), namely RGS6, RGS7, RGS9 and RGS11, are expressed in the retina along with its binding partner Gβ5. The RGS9-1 isoform is expressed only in retinal photoreceptors and rate-limits the recovery of rod phototransduction by acting as a member of the transducin GAP complex (RGS9-1/Gβ5L/R9AP). The Gβ5L isoform is also only expressed in retinal photoreceptors and acts by stabilizing the GAP complex. The Gβ5S isoform differs from Gβ5L by the absence of exon 1 due to alternative splicing and is expressed in many other retinal cells. Gβ5L is barely detectable in RGS9-/- …

Signals Delivered By Interleukin-7 Regulate The Activities Of Bim And Jund In T Lymphocytes, Shannon Moore Ruppert

Electronic Theses and Dissertations

Interleukin-7 (IL-7) is an essential cytokine for lymphocyte growth that has the potential for promoting proliferation and survival. While the survival and proliferative functions of IL-7 are well established, the identities of IL-7 signaling components in pathways other than JAK/STAT, that accomplish these tasks remain poorly defined. To this end, we used IL-7 dependent T-cells to examine those components necessary for cell growth and survival. Our studies revealed two novel signal transducers of the IL-7 growth signal: BimL and JunD. IL-7 promoted the activity of JNK (Jun N-terminal Kinase), and that JNK, in turn, drove the expression of JunD, a …

Expression Of Lipase From Mycobacterium Tuberculosis In Nicotiana Tobacum And Lactuca Sativa Chloroplasts, Bethany Lloyd

Electronic Theses and Dissertations

Tuberculosis (TB), caused by the bacterium Mycobacterium tuberculosis (M. tuberculosis), is a global threat and the leading cause of death among individuals infected with HIV. TB treatment requires multi-drug cocktails, due to the increasing rates of drug resistance of the bacterium. With multi-drug cocktails, strains have been documented to be resistant to all major drugs in the fight against TB. Since the strains are drug resistant, it calls for an increasing need for vaccine and treatment development for the purpose of preventing and managing the disease. The most widely distributed vaccine against TB is Bacillus Calmette-Gue´rin (BCG). Apart from being …

Cerium Oxide Nanoparticles Act As A Unique Catalyst And Scavenge Nitric Oxide And Peroxynitrite And Decrease Rns In Vitro And In Vivo, Janet Dowding

Electronic Theses and Dissertations

Cerium oxide nanoparticles (CeO2 NPs)(nanoceria) have been shown to possess a substantial oxygen storage capacity via the interchangeable surface reduction and oxidation of cerium atoms, cycling between the Ce4+ and Ce3+ redox states. Reduction of Ce4+ to Ce3+ causes oxygen vacancies or defects on the surface of the crystalline lattice structure of the particles, generating a cage for redox reactions to occur. The study of the chemical and biological properties of CeO2 NPs has expanded recently, and the methods used to synthesize these materials are also quite diverse. This has led to a plethora of studies describing various preparations of …

Manipulation Of Host Signal Transduction Pathways And Cytoskeleton Functions By Invasive Bacterium Listeria Monocytogenes And Chlamydia Trachomatis, Shahanawaz Jiwani

Electronic Theses and Dissertations

Infectious disease remains one of the leading causes of morbidity and mortality worldwide. Many bacteria that cause disease have the capacity to enter into eukaryotic cells such as epithelial cells and tissue macrophages. Gaining access into the intracellular environment is one of the most critical steps in their survival and/or in pathogenesis. The entry mechanisms employed by these organisms vary considerably, but most mechanisms involve sabotaging and manipulating host cell functions. Invasion of epithelial cells involves triggering host signal transduction mechanisms to induce cytoskeleton rearrangement, thereby facilitating bacterial uptake. My work focuses on understanding the molecular mechanisms employed by bacterial …

Defective Dynamics Of Mitochondria In Amyotrophic Lateral Sclerosis And Huntington's Disease, Wenjun Song

Electronic Theses and Dissertations

Mitochondria play important roles in neuronal function and survival, including ATP production, Ca2+ buffering, and apoptosis. Mitochondrial dysfunction is a common event in the pathogenesis of neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS) and Huntington's disease (HD); however, what causes the mitochondrial dysfunction remains unclear. Mitochondrial fission is mediated by dynamin-related protein 1 (DRP1) and fusion by mitofusin 1/2 (MFN1/2) and optic atrophy 1 (OPA1), which are essential for mitochondrial function. Mutations in the mitochondrial fission and fusion machinery lead to neurodegeneration. Thus, whether defective mitochondrial dynamics participates in ALS and HD requires further investigation. ALS is a fatal …

Novel Immunogens Of Cellular Immunity Revealed Using In Vitro Human Cell-Based Approach, Brian Schanen

Electronic Theses and Dissertations

In the last 150 years, tremendous headway has been made in our understanding of the human immune system. Pioneers in the field such as Paul Ehrlich, Elie Metchnikoff, Louis Pasteur, Robert Koch and Walter Reed carried out seminal studies that established the groundwork for our understanding of humoral and cellular immunity in humans. However, this direct line of evidence into human immunology was diverted in the mid-20th century with the adoption of a model which allowed for investigators to use a reductionist-approach with the promise to resolve immunity at a molecular level. This revolutionary model was the scientific commercialization of …

Integrated Assessment Of Anthropogenic, Climate, And Policy Induced Changes Of Phosphorus Export In The United States Laurentian Great Lakes Watersheds, Meredith Ballard Labeau

Dissertations, Master's Theses and Master's Reports - Open

Anthropogenic activities have increased phosphorus (P) loading in tributaries to the Laurentian Great Lakes resulting in eutrophication in small bays to most notably, Lake Erie. Changes to surface water quality from P loading have resulted in billions of dollars in damage and threaten the health of the world’s largest freshwater resource. To understand the factors affecting P delivery with projected increasing urban lands and biofuels expansion, two spatially explicit models were coupled. The coupled models predict that the majority of the basin will experience a significant increase in urban area P sources while the agriculture intensity and forest sources of …

Biophysics, Environmental Stochasticity, And The Evolution Of Thermal Safety Margins In Intertidal Limpets, W. Wesley Dowd, M. W. Denny

Biology Faculty Works

As the air temperature of the Earth rises, ecological relationships within a community might shift, in part due to differences in the thermal physiology of species. Prediction of these shifts – an urgent task for ecologists – will be complicated if thermal tolerance itself can rapidly evolve. Here, we employ a mechanistic approach to predict the potential for rapid evolution of thermal tolerance in the intertidal limpet Lottia gigantea. Using biophysical principles to predict body temperature as a function of the state of the environment, and an environmental bootstrap procedure to predict how the environment fluctuates through time, we create …

Cell Cycle Arrest Associated With Anoxia-Induced Quiescence, Anoxic Preconditioning, And Embryonic Diapause In Embryos Of The Annual Killifish Austrofundulus Limnaeus, Camie Lynn Meller, Robert Meller, Roger P. Simon, Kristin M. Culpepper, Jason E. Podrabsky

Biology Faculty Publications and Presentations

Embryos of the annual killifish Austrofundulus limnaeus can enter into dormancy associated with diapause and anoxia-induced quiescence. Dormant embryos are composed primarily of cells arrested in the G1/G0 phase of the cell cycle based on flow cytometry analysis of DNA content. In fact, most cells in developing embryos contain only a diploid complement of DNA, with very few cells found in the S, G2, or M phases of the cell cycle. Diapause II embryos appear to be in a G0-like state with low levels of cyclin D1 and p53. However, the active form of pAKT is high during diapause II. …

Investigations Into Modulation Of Brain Oxidative Stress By Various Interventions, Jessica Lynn Harris

Theses and Dissertations--Chemistry

In this thesis study we examined glycogen synthase kinase-3β (GSK-3β) and its effects over Nrf2 and Pin 1 as it relates to Alzheimer’s disease (AD). AD is a neurodegenerative disease characterized by a prolonged high oxidative environment. Transcription factor Nrf2 is vital in the brain’s defense against oxidative insults through its up-regulation of over 100 antioxidants. Depletion of the brain’s antioxidant defense system results in intolerance to an oxidative environment, contributing to the progression of AD. The regulatory Pin 1 protein promotes cellular homeostasis, and when down-regulated results in increased deposits of neurofibrillary tangles (NFTs) and amyloid-β (Aβ) plaques, the …

Bifunctional Bisphosphonates For Delivering Biomolecules To Bone, Jivan N. Yewle

Theses and Dissertations--Chemistry

Active targeting with controlled delivery of therapeutic agents to bone is an ideal approach for treatment of several bone diseases. Since bisphosphonates (BPs) are known to have high affinity to bone mineral and are being widely used in treatment of osteoporosis, they are well-suited for drug targeting to bone. For this purpose, bifunctional hydrazine-bisphosphonates (HBPs) with spacers of various lengths and lipophilicity were synthesized and studied. Crystal growth inhibition assays demonstrated that the HBPs with shorter spacers bound more strongly to bone mineral, hydroxyapatite (HA), than did alendronate. HBPs were also demonstrated to be non-toxic to MC3T3-E1 pre-osteoblasts. The targeted …

Design, Synthesis, And Anticancer Activity Of Ruthenium Complexes, Brock S. Howerton

Theses and Dissertations--Chemistry

Ruthenium complexes show promise as light activated photodynamic therapy (PDT) prodrugs. Strained octahedral complexes were synthesized that produce a cytotoxic species upon light activation. pUC19 DNA damage in vitro experiments were carried out to determine the type of damage observed. In vivo cell experiments were carried out on the non-small lung cancer A549 cell line to determine the phototherapeutic window of the synthesized complexes. One mechanism of drug resistance via elevated levels of glutathione was addressed through in vitro binding studies carried out with UV-Vis spectroscopy and in vivo glutathione titrations in the A549 cell line. Several complexes were shown …

Examination Of The Neurochemical Mechanisms That Mediate Nicotine Withdrawal In Adolescent And Adult Rats, Luis Alberto Natividad

Open Access Theses & Dissertations

Introduction: The mechanisms that mediate nicotine withdrawal are presently unclear and age group differences in the neurochemical effects of withdrawal have been largely unexplored. Previous studies in our laboratory demonstrated that adult rats display a decrease in extracellular levels of dopamine in the nucleus accumbens (NAcc) during nicotine withdrawal and this decrease is reduced in adolescent rats (Natividad et al., 2010). The goal of this dissertation was to examine whether these age group differences in dopamine during withdrawal are mediated via excitatory and inhibitory mechanisms that modulate dopamine in the cell body region of the ventral tegmental area (VTA). Methods: …

Further Characterization Of The Nodamura Virus Rna2 3'-Terminal Stem Loop Structure And Its Role In Viral Rna Replication, Joshua Frederick

Open Access Theses & Dissertations

Nodamura virus (NoV) is an excellent model system for studying many aspects of the life cycle of positive-strand RNA viruses. NoV's simple genome organization and ability to replicate its genome to high levels in a wide variety of host cells, including plants, yeast, insects and mammals (Ball et al., 1992; Garzon et al., 1978, 1990; Murphy et al., 1970; Price et al., 2005; Scherer & Hurlbut, 1967; Selling et al., 1990; reviewed by Ball & Johnson, 1998), make it a prime model system for studying the basic mechanisms of viral RNA replication. Other positive-strand RNA viruses, including those that are …

The Effects Of Extended Access To Methamphetamine Self-Administration On Dopaminergic Markers In The Striatum, Joe Luevano

Open Access Theses & Dissertations

Methamphetamine (METH) abuse is a persistent problem in the U.S. and abroad. Escalation of METH use among independent users occurs for a variety of physiological and psychological reasons. Methamphetamine dependence may be attributed to the rewarding effect of this drug via the dopaminergic systems of the central nervous system (CNS). The presence of METH in the CNS increases synaptic release of dopamine. This increase in dopaminergic neurotransmission is thought to be directly attributed to the rewarding effects of METH. Following METH use, compensatory changes have been found to occur in the dopaminergic system during various periods of abstinence. It is …

Arachidonic Acid Signaling In Invasive And Non-Invasive Breast Cancer Cells, Debarshi Roy

Open Access Theses & Dissertations

Breast cancer is the second largest cause of cancer-related deaths in women all over the world. Epidemiological studies suggest that the consumption of high-fat diets can promote the incidence of breast cancers in both developed and developing countries. In particular, the lipid-rich diet contains arachidonic acid (AA, a C20:4 polyunsaturated fatty acid), which has been shown to be associated with tumor formation in breast tissues. Nevertheless, the actual mechanism by which AA induces the metastatic transformation and malignancy is not well understood. The goal of my dissertation, therefore, is to identify the molecules and unravel the pathways that participate in …

Determination Of An Interaction Between Nipped B-Like Protein And Mll, Adam Robert Marek

Master's Theses

The Mixed Lineage Leukemia (MLL) protein serves as a positive transcriptional regulator during hematopoietic and embryonic development. The MLL gene can undergo chromosomal translocations producing leukemia-causing fusions that retain the MLL amino-terminus, including the repression domain. A recent yeast two-hybrid screening used the MLL repression domain as bait and yielded nine positive clones of Nipped B-like (NIPBL).

NIPBL is a crucial member of the cohesin complex, which functions in the segregation of sister chromatids during cell division. However, recent evidence suggests the cohesin complex can also function as a transcriptional regulator.

In this study, we wanted to confirm this interaction …

Cloning Of The Gene, Purification As Recombinant Protein And Functional Characterization Of Leishmania Mexicana Cytochrome B5 Reductase, Ala Azhari

USF Tampa Graduate Theses and Dissertations

Leishmania are protozoan parasites that are transmitted by a sand fly vector. These parasites affect not only humans but also wild animals including domestic dogs and rodents, which form an additional challenge and public health problem to control the disease. Leishmaniasis is an important disease with worldwide distribution, including Saudi Arabia, the Middle East, and other tropical and subtropical areas around the world. Due to the expansion of irrigation and agricultural activities, more exposure to sand fly occurs, which leads to the expansion of leishmaniasis infections as newly emerging disease.

Emerging drug resistance in leishmaniasis is an additional problem, contributed …

Aberrant Sialylation Alters Cardiac Electrical Signaling, Andrew Ednie

USF Tampa Graduate Theses and Dissertations

In the heart, electrical signaling is responsible for its rhythmicity and is necessary to initiate muscle contraction. The net electrical activity in a cardiac myocyte during a contraction cycle is observed as the action potential (AP), which describes a change in membrane potential as a function of time. In ventricular cardiac myocytes, voltage-gated sodium channels (Nav) and voltage-gated potassium channels (Kv) play antagonistic roles in shaping the AP with the former initiating membrane depolarization and the latter repolarizing it. Functional changes in the primary cardiac Nav isoform, Nav 1.5, or any one of the many Kv isoforms expressed in the …

Development Of An In-Cell Förster Resonance Energy Transfer Technique To Study Protein Structure Inside Living Cells, Victoria Lynn Murray

Wayne State University Dissertations

The goal of my thesis is to develop an in-cell fluorescence technique that allows for measurement of the distances between fluorescence acceptors and donors within a protein or between two proteins inside the correct intracellular compartment of living cells. The successful achievement of this goal will allow us to obtain high-resolution structural information from a protein, one key step towards high-resolution structural biology of proteins inside the living cell.

To achieve this goal, we will apply the fluorescence resonance energy transfer (FRET) technique to the specifically labeled proteins inside the cells. Our rationale is to specifically label the protein(s) of …

The Speed Of Sound And Attenuation Of An Iec Agar-Based Tissue-Mimicking Material For High Frequency Ultrasound Applications, Chao Sun, Stephen Pye, Jacinta Browne, Anna Janeczko, Bill Ellis, Mairead Butler, Vassilis Sboros, Adrain Thomson, Mark Brewin, Charles Earnshaw, Carmel Moran

Articles

No abstract provided.

Hdm2 Small-Molecule Inhibitors For Therapeutic Intervention In B-Cell Lymphoma, Angela Sosin

Wayne State University Dissertations

Lymphomas frequently retain wild-type (wt) p53 function but overexpress HDM2, compromising p53 activity. Therefore, lymphoma is a suitable model for studying therapeutic value of disrupting HDM2-p53 association by small-molecule inhibitors (SMIs). HDM2 SMIs have been developed and are currently under various stages of preclinical and clinical investigation. This study examined various molecular mechanisms associated and biological effects of two different classes of HDM2 SMIs: the spiro-oxindoles (MI-219) and cis-imidazoline (Nutlin-3) in lymphoma cell lines and patient-derived B-lymphoma cells. Surprisingly, results revealed significant quantitative and qualitative differences between these two agents. At the molecular level, effect of Nutlin-3 was generally more …

Bacterial Host Interactions In Cystic Fibrosis, Máire Callaghan, Siobhan Mcclean

Articles

Chronic infection is a hallmark of cystic fibrosis (CF) and the main contributor to morbidity. Microbial infection in CF is complex, due to the number of different species that colonise the CF lung. Their colonisation is facilitated by a host response that is impaired or compromised by highly viscous mucous, zones of hypoxia and the lack of the cystic fibrosis transmembrane regulator (CFTR). Successful dominant CF pathogens combine an effective arsenal to establish infection and counter-attack the host response, together with an ability to adapt readily to an unfavourable environment. Hypermutability is common among CF pathogens facilitating adaptation and as …