Biochemistry, Biophysics, and Structural Biology Commons^™

Retention And Loss Of Rna Interference Pathways In Trypanosomatid Protozoans, Lon-Fye Lye, Katherine Owens, Huafang Shi, Silvane M. F. Murta, Ana Carolina Vieira, Salvatore J. Turco, Christian Tschudi, Elisabetta Ullu, Stephen M. Beverley

Molecular and Cellular Biochemistry Faculty Publications

RNA interference (RNAi) pathways are widespread in metaozoans but the genes required show variable occurrence or activity in eukaryotic microbes, including many pathogens. While some Leishmania lack RNAi activity and Argonaute or Dicer genes, we show that Leishmania braziliensis and other species within the Leishmania subgenus Viannia elaborate active RNAi machinery. Strong attenuation of expression from a variety of reporter and endogenous genes was seen. As expected, RNAi knockdowns of the sole Argonaute gene implicated this protein in RNAi. The potential for functional genetics was established by testing RNAi knockdown lines lacking the paraflagellar rod, a key component of the …

Characterization Of The Role Of Nicotine And Delta 9-Thc In Modulation Of Neuroinflammation, Jared C. Ehrhart

USF Tampa Graduate Theses and Dissertations

Neuroinflammation is a major driving force in the progression of neurodegenerative disorders. Nicotinic acetylcholine receptors, as well as cannabinoid CB2 receptors, have been shown to have strong anti-inflammatory properties when activated. These effects are shown, in vivo, to be a result of stimulation of α7 nAChRs and CB2 cannabinoid receptors. Microglia cells, an immune cell in the brain, are shown to express both of these receptor subtypes. The studies detailed herein, investigated the ability of two compounds, nicotine and Δ9-THC, in modulation of inflammatory processes. Stimulation of these receptors on microglia using nicotine and Δ9-THC blocked the activation of these …

Adenovirus-5 Infection Affects Lipid Metabolism In Hepatic And Adipose Tissues, Marianna Sukholutsky

Theses and Dissertations

Our recent studies have shown a link between Adenovirus-5 (Ad-5) and elevated lipids, which prompted the hypothesis that Ad-5 infection might augment hepatic and/or adipose tissue lipid metabolism. To test our hypothesis, mice were infected with Ad-5 and screened for changes in lipogenesis and plasma markers associated with the metabolic syndrome. We observed increased expression of sterol regulatory element binding protein 1 (SREBP-1) in infected liver tissues, but not in adipose tissues and this correlated with elevated plasma and hepatic triglyceride levels. Elevated expression of adiponectin was seen in Ad-5 infected adipose tissues and this correlated with phosphorylated AMPK in …

(1e,3e)-1,4-Bis(4-Methoxyphenyl)Buta1,3-Diene, Gopinathan Narayan, Nigam Rath, Suresh Das

Chemistry & Biochemistry Faculty Works

The title compound, C18H18O2, which exhibits blue emission in the solid state, is an inter­mediate in the preparation of liquid crystals and polymers. The mol­ecule is located on an inversion centre. In the crystal, mol­ecules are arranged in a herringbone motif.

Molecular Mechanism Of Agc Kinases In Human Malignant, Shaokun Shu

USF Tampa Graduate Theses and Dissertations

The maintenance of normal cell function and tissue homeostasis is dependent on the precise regulation of multiple signaling pathways that control cellular decisions to either proliferate, differentiate, arrest cell growth, or initiate programmed cell death (apoptosis). Cancer arises when clones of mutated cells escape this balance and proliferate inappropriately without compensatory apoptosis. Deregulated cell growth occurs as a result of perturbed signal transduction that modulates or alters cellular behavior or function to keep the critical balance between the rate of cell-cycle progression (cell division) and cell growth (cell mass) on one hand, and programmed cell death (apoptosis, autophagy) on the …

Functionalization Of Diamond And Its Application In High Performance Liquid Chromatography And Solid Phase Extraction, Gaurav Saini

Theses and Dissertations

The primary focus of my work was to chemically functionalize diamond as normal and reversed phases for solid phase extraction (SPE) and high performance liquid chromatography (HPLC). Diamond was functionalized with -NH2 groups via self-limiting adsorption of an amine-containing polymer, polyallylamine (PAAm), onto oxidized diamond particles. The chemical stability of these particles was improved by thermal curing or chemical crosslinking with 1,2,5,6-diepoxycyclooctane. The reversed phase material for SPE was synthesized by reacting amine-functionalized diamond particles with long chain alkyl and a perfluorinated isocyanate. Unlike commercially available silica, functionalized diamond particles were stable under extreme pH conditions (pH 0-14). Functionalized diamond …

Scoring Function To Predict Solubility Mutagenesis, Ye Tian, Christopher Deutsch, Bala Krishnamoorthy

Chemistry Faculty Publications and Presentations

Background: Mutagenesis is commonly used to engineer proteins with desirable properties not present in the wild type (WT) protein, such as increased or decreased stability, reactivity, or solubility. Experimentalists often have to choose a small subset of mutations from a large number of candidates to obtain the desired change, and computational techniques are invaluable to make the choices. While several such methods have been proposed to predict stability and reactivity mutagenesis, solubility has not received much attention. Results: We use concepts from computational geometry to define a three body scoring function that predicts the change in protein solubility due to …

Solving The Low Dimensional Smoluchowski Equation With A Singular Value Basis Set, Gregory E. Scott, Martin Gruebele

Chemistry and Biochemistry

Reaction kinetics on free energy surfaces with small activation barriers can be computed directly with the Smoluchowski equation. The procedure is computationally expensive even in a few dimensions. We present a propagation method that considerably reduces computational time for a particular class of problems: when the free energy surface suddenly switches by a small amount, and the probability distribution relaxes to a new equilibrium value. This case describes relaxation experiments. To achieve efficient solution, we expand the density matrix in a basis set obtained by singular value decomposition of equilibrium density matrices. Grid size during propagation is reduced from (100–1000) …

The Expression Patterns Of Minor Fibrillar Collagens During Development In Zebrafish, Ming Fang, Jason S. Adams, B. Lane Memahhan, Raquel J. Brown, Julia Oxford

Biomedical Research Institute Publications and Presentations

Minor fibrillar collagens are recognized as the organizers and nucleators during collagen fibrillogenesis but likely serve additional functions. The minor fibrillar collagens include collagens type V and type XI. Mutations of collagen type V and XI can cause Ehlers Danlos, Stickler's, and Marshall's syndromes in human. We have characterized the spatiotemporal expression patterns of Col11a1, Col11a2, Col5a1 as well as Col5a3 in zebrafish embryos by in situ hybridization. Col5a1 is expressed in developing somites, neural crest, the head mesenchyme, developing cranial cartilage, pharyngeal arches and vertebrae. Col5a3 is detected in the notochord, mesenchyme cells in the eyes and lens. Both …

Biocompatibility Of Synthetic Poly(Ester Urethane)/Polyhedral Oligomeric Silsesquioxane Matrices With Embryonic Stem Cell Proliferation And Differentiation, Yan-Lin Guo, Wenshou Wang, Joshua U. Otaigbe

Faculty Publications

Incorporation of polyhedral oligomeric silsesquioxanes (POSS) into poly(ester urethanes) (PEU) as a building block results in a PEU/POSS hybrid polymer with increased mechanical strength and thermostability. An attractive feature of the new polymer is that it forms a porous matrix when cast in the form of a thin film, making it potentially useful in tissue engineering. In this study, we present detailed microscopic analysis of the PEU/POSS matrix and demonstrate its biocompatibility with cell culture. The PEU/POSS polymer forms a continuous porous matrix with open pores and interconnected grooves. From SEM image analysis, it is calculated that there are about …

Synthesis And Structural Characterization Of The Peptide Epitope Of The Ovarian Cancer Biomarker Ca125 (Muc16), Rebecca J. Whelan, Zach T. Berman, Lee J. Moore, Kathleen E. Knudson

Faculty & Staff Scholarship

A highly conserved region of 21 amino acids flanked by cysteine residues, contained within a larger repeated domain, has been proposed to be the antibody-binding site in the ovarian cancer biomarker CA125 (MUC16). In this study solid-phase peptide synthesis with Fmoc protection chemistry was used to assemble a 21-mer peptide corresponding to the most frequently occurring antibody binding sequence in CA125. Potentially significant sequence variants were also synthesized. Peptide secondary structure was investigated using Fourier transform infrared spectroscopy, revealing the consensus sequence peptide to be largely unstructured at physiological pH whether the cysteine residues were reduced or were oxidized to …

Structure Of The Cholera Toxin Secretion Channel In Its Closed State, Steve Reichow, Konstantin V. Korotkov, Wim Gj Hol, Tamir Gonen

Chemistry Faculty Publications and Presentations

The type II secretion system (T2SS) is a macromolecular complex spanning the inner and outer membranes of Gram-negative bacteria. Remarkably, the T2SS secretes folded proteins including multimeric assemblies like cholera toxin and heat-labile enterotoxin from Vibrio cholerae and enterotoxigenic Escherichia coli, respectively. The major outer membrane T2SS protein is the “secretin” GspD. Electron cryomicroscopy reconstruction of the V. cholerae secretin at 19 Å resolution reveals a dodecameric structure reminiscent of a barrel with a large channel at its center that appears to contain a closed periplasmic gate. The GspD periplasmic domain forms a vestibule with a conserved constriction, and binds …

Cooperative Interaction Of Transcription Termination Factors With The Rna Polymerase Ii C-Terminal Domain, Bradley M. Lunde, Steve Reichow, Minkyu Kim, Hyunsuk Suh, Thomas C. Leeper, Fan Yang, Hannes Mutschler, Stephen Buratowski, Anton Meinhart, Gabriele Varani

Chemistry Faculty Publications and Presentations

Phosphorylation of the C-terminal domain of RNA polymerase II controls the co-transcriptional assembly of RNA processing and transcription factors. Recruitment relies on conserved CTDinteracting domains that recognize different CTD phosphoisoforms during the transcription cycle, but the molecular basis for their specificity remains unclear. We show that the CTD-interacting domains of two transcription termination factors, Rtt103 and Pcf11, achieve high affinity and specificity both by specifically recognizing the phosphorylated CTD and by cooperatively binding to neighboring CTD repeats. Single amino acid mutations at the protein-protein interface abolish cooperativity and affect recruitment at the 3′-end processing site in vivo. We suggest that …

The Human In 3d: Advanced Morphometric Analysis Of High-Resolution Anatomically Accurate Computed Models, Summer J. Decker

USF Tampa Graduate Theses and Dissertations

Computed virtual models of anatomical structures are proving to be of increasing value in clinical medicine, education and research. With a variety of fields focused on craniofacial and pelvic anatomy there is a need for accurate anatomical models. Recent technological advancements in computer and medical imaging technologies have provided the tools necessary to develop three-dimensional (3D) functional models of human anatomy for use in medicine (surgical planning and education), forensics and engineering (computer-aided design (CAD) and finite element analysis).

Traditionally caliper methodologies are used in the quantitative analysis of human anatomy. In order for experts in anatomy and morphometrics to …

Phosphorus Uptake In Rhodomonas Salina (Wislouch) And Its Effect On Allocation And Elimination In Acartia Tonsa (Dana), Danna Palladino

OES Theses and Dissertations

Phosphorus is a key element in important biochemical compounds, such as RNA and phospholipids, and can become limiting in a variety of marine systems. The uptake of phosphorus into biochemical fractions (protein, low molecular weight (LMW) compounds, lipid, polysaccharide and nucleic acid) in Acartia tonsa fed ³³P -labeled Rhodomonas salina was examined. R. salina was cultured on two variations of one media that in one case contained phosphorus in balance and the other out of balance with relation to other standard f/2 components. The P-balanced (PB) media had a N:P ratio of 24.5, which is higher than that found …

Characterization Of Metastasis-Associated Cell Surface Glycoproteins In Prostate Cancer, Lifang Yang

Theses and Dissertations in Biomedical Sciences

Prostate cancer (PCa) is a major health problem in males in the United States. Its lethality is mostly attributed to the primary tumor metastasizing to distant sites that are highly resistant to conventional therapies. Serum Prostate Specific Antigen (PSA) is the only protein biomarker used in clinic for prediction of prostate cancer recurrence following local therapies. Nonetheless, PSA lacks the ability to predict the behavior of an individual tumor in an individual patient. Therefore, development of reliable biomarkers for detection of metastatic potential in primary tumors, as well as discovery of new therapeutic targets, is in a great need for …

The Biogeochemistry Of Submerged Coastal Karst Features In West Central Florida, Keith Michael Garman

USF Tampa Graduate Theses and Dissertations

West Central Florida is a complex karst environment with numerous sinkholes, springs, and submerged cave systems. Many of these karst features are anchialine, located within the subterranean estuary where freshwater and saltwater mix. Water quality data and/or fauna data were obtained from twenty-one karst features and their associated cave systems. The anchialine karst environment of the study area has a wide range of habitats with measured salinities ranging from freshwater at <0.2 ppt to sulfidic, hypersaline water at 38.5 ppt and measured pH readings ranging from 6.39 in water impacted by sulfur oxidizing bacteria to 10.3 in an isolated room of a cave. Stygobitic crustaceans were identified in conduits extending beneath the Gulf of Mexico supporting the hypotheses that freshwater crustaceans could survive higher sea levels in freshwater conduits beneath saltwater. The fauna associated with the anchialine cave systems included Sabellidae and Polychaeta worms, hydroids, cnidarians and hydrobiid snails. Jewfish Sink, like other anaerobic marine basins that were submarine springs, has four zones: oxic zone, transition zone, upper anoxic zone and anoxic bottom water. The upper zones have seasonal water quality variations from winter cooling and sinking of surface water and changes in the microbial communities. Activity of sulfate reducing bacteria is carbon limited in the anoxic zones, where sulfate reduction is the major metabolic process, and primary production is phosphate limited in the oxic zones. Organic input from the Gulf of Mexico drives the bacterial anaerobic ecosystem, resulting in a “sulfide pump”, in which sulfide percolates upward removing oxygen from the overlying sediment.

Reasons For Reducing Breastfeeding Duration Among Chinese Mothers In Ireland:A Qualitative Study, Qianling Zhou

Other resources

Introduction: The Ireland Chinese Mother Survey has revealed a remarkable decline of breastfeeding duration among Chinese mothers who gave birth in Ireland [1]. This qualitative study was undertaken to explore the reasons for such a decrease and to determine the influence of residency in Ireland on breastfeeding practices among immigrant Chinese. Methods: Purposive sampling together with the snowball technique was used to recruit immigrant Chinese women who have given birth in Ireland. Seven focus groups were conducted and thematic content analysis was performed to analyse the data. Results: Decline in breastfeeding duration was not mainly due to the formula feeding …

The Mir-15/107 Group Of Microrna Genes: Evolutionary Biology, Cellular Functions, And Roles In Human Diseases, John R. Finnerty, Wang-Xia Wang, Sébastien S. Hébert, Bernard R. Wilfred, Guogen Mao, Peter T. Nelson

Pathology and Laboratory Medicine Faculty Publications

The miR-15/107 group of microRNA (miRNA) gene is increasingly appreciated to serve key functions in humans. These miRNAs regulate gene expression involved in cell division, metabolism, stress response, and angiogenesis in vertebrate species. The miR-15/107 group has also been implicated in human cancers, cardiovascular disease and neurodegenerative disease, including Alzheimer's disease. Here we provide an overview of the following: (1) the evolution of miR-15/107 group member genes; (2) the expression levels of miRNAs in mammalian tissues; (3) evidence for overlapping gene-regulatory functions by different miRNAs; (4) the normal biochemical pathways regulated by miR-15/107 group miRNAs; and (5) the roles played …

The Regulation Of The Alternative Splicing Of Caspase 9, Rachel Goehe

Theses and Dissertations

The pro-apoptotic, caspase 9a, and the anti-apoptotic, caspase 9b, are derived from the caspase 9 gene by alternative splicing. This study demonstrates that the alternative splicing of caspase 9 is dysregulated in a large percentage of non-small cell lung cancer (NSCLC) tumors of the adenocarcinoma type. Furthermore, modulation of the levels of splice variants of caspase 9 had dramatic effects on the anchorage-independent growth and tumorigenic capacity of NSCLC cells. Due to these findings, the molecular mechanisms regulating the post-transcriptional processing of caspase 9 were therefore examined and an exonic splicing silencer (ESS) regulating the pre-mRNA processing of caspase 9 …

Physical Interaction Between Vivid And White Collar Complex Regulates Photoadaptation In Neurospora, Chen-Hui H. Chen, Bradley S. Demay, Amy S. Gladfelter, Jay Dunlap, Jennifer J. Loros

Dartmouth Scholarship

Photoadaptation, the ability to attenuate a light response on prolonged light exposure while remaining sensitive to escalating changes in light intensity, is essential for organisms to decipher time information appropriately, yet the underlying molecular mechanisms are poorly understood. In Neurospora crassa, VIVID (VVD), a small LOV domain containing blue-light photoreceptor protein, affects photoadaptation for most if not all light-responsive genes. We report that there is a physical interaction between VVD and the white collar complex (WCC), the primary blue-light photoreceptor and the transcription factor complex that initiates light-regulated transcriptional responses in Neurospora. Using two previously characterized VVD mutants, we show …

The Terebridae And Teretoxins: Combining Phylogeny And Anatomy For Concerted Discovery Of Bioactive Compounds, Nicolas Puillandre, Mandë Holford

Publications and Research

The Conoidea superfamily, comprised of cone snails, terebrids, and turrids, is an exceptionally promising group for the discovery of natural peptide toxins. The potential of conoidean toxins has been realized with the distribution of the first Conus (cone snail) drug, Prialt (ziconotide), an analgesic used to alleviate chronic pain in HIV and cancer patients. Cone snail toxins (conotoxins) are highly variable, a consequence of a high mutation rate associated to duplication events and positive selection. As Conus and terebrids diverged in the early Paleocene, the toxins from terebrids (teretoxins) may demonstrate highly divergent and unique functionalities. Recent analyses of the …

Strategies For The Elimination Of Matrix Effects In The Lc-Ms/Ms Analysis Of The Lipophilic Toxins Okadaic Acid And Azaspiracid-1 In Molluscan Shellfish, Jane Kilcoyne, Elie Fux

Articles

Considerable efforts are being made worldwide to replace in vivo assays with instrumental methods of analysis for the monitoring of marine biotoxins in shellfish. Analysis of these compounds by the preferred technique of LC-MS/MS is challenged by matrix effects associated with shellfish tissue components. In methods validation, assessment of matrix interferences is imperative to ensure the accuracy of analytical results. We evaluated matrix interferences in the analysis of okadaic acid (OA) and azaspiracid 1 (AZA1) in mollucscan shellfish by using a conventional acidic method on electrospray triple stage quadrapole (TSQ) and hybrid quadrupole time of flight (QToF) instruments, with matrix …

Aromatic Acid Metabolites Of Escherichia Coli K-12 Can Induce The Marrab Operon, Lon Chubiz, Christopher Rao

Biology Department Faculty Works

MarR is a key regulator of the marRAB operon involved in antibiotic resistance and solvent stress tolerance in Escherichia coli. We show that two metabolic intermediates, 2,3-dihydroxybenzoate and anthranilate, involved in enterobactin and tryptophan biosynthesis, respectively, can activate marRAB transcription. We also found that a third intermediate involved in ubiquinone biosynthesis, 4-hydroxybenzoate, activates marRAB transcription in the absence of TolC. Of the three, however, only 2,3-dihydroxybenzoate directly binds MarR and affects its activity.

Specificity Determinants Of Arma, A Ribosomal Rna Methyltransferase That Confers Antibiotic Resistance, Tamara Zarubica

Theses and Dissertations

Bacterial resistance to 4,6-type aminoglycoside antibiotics, which target the 30S ribosomal subunit, has been traced to the arm/rmt family of rRNA methyltransferases. These plasmid-encoded enzymes transfer a methyl group from S-adenosylmethionine to N7 of the buried G1405 in the aminoglycoside binding site of 16S rRNA in the 30S ribosomal subunit. Neither 16S rRNA alone nor intact 70S ribosome is an efficient substrate for armA methyltransferase. To more fully characterize this family of enzymes, xiii we have investigated the substrate requirements of ArmA. We determined the Mg2+ dependence of ArmA activity and found that the enzyme could recognize both translationally active …

Axial Temperature Gradients In Gas Chromatography, Jesse Alberto Contreras

Theses and Dissertations

The easiest and most effective way to influence the separation process in gas chromatography (GC) is achieved by controlling the temperature of the chromatographic column. In conventional GC, the temperature along the length of the column is constant at any given time, T(t). In my research, I investigated the effects of temperature gradients on GC separations as a function of time and position, T(t,x), along the column. This separation mode is called thermal gradient GC (TGGC). The research reported in this dissertation highlights the fundamental principles of axial temperature gradients and the separation potential of the TGGC technique. These goals …

Maternal Health Behaviours During Pregnancy And Associations With Infant Feeding Factors, Roslyn Tarrant, Katherine Younger, Margaret Sheridan-Pereira, John Kearney

Other resources

Maternal health behaviours during pregnancy and associations with postnatal infant feeding practices

Objectives: To examine modifiable maternal health behaviours during pregnancy, including folic acid use peri-conceptually, smoking and alcohol consumption patterns, and relate these factors to maternal postnatal infant feeding practices.

Materials and Methods: A prospective observational study involved the recruitment of 539 pregnant women from a Dublin maternity hospital. Following application of exclusion criteria, 450 mothers were followed up at 6 weeks and 6 months postpartum. Quantitative data on folic acid use, as well as mothers’ smoking and alcohol consumption patterns during pregnancy were collected from the antenatal questionnaire. …

How To Be Successful In Exclusive Breastfeeding For Four To Six Months:Experiences Of Chinese Mothers In Ireland, Qianling Zhou

Other resources

No abstract provided.

Negative Dielectrophoretic Capture Of Bacterial Spores In Food Matrices, Mehti Koklu, Seungkyung Park, Suresh D. Pillai, Ali Beskok

Mechanical & Aerospace Engineering Faculty Publications

A microfluidic device with planar square electrodes is developed for capturing particles from high conductivity media using negative dielectrophoresis (n-DEP). Specifically, Bacillus subtilis and Clostridium sporogenes spores, and polystyrene particles are tested in NaCl solution (0.05 and 0.225 S/m), apple juice (0.225 S/m), and milk (0.525 S/m). Depending on the conductivity of the medium, the Joule heating produces electrothermal flow (ETF), which continuously circulates and transports the particles to the DEP capture sites. Combination of the ETF and n-DEP results in different particle capture efficiencies as a function of the conductivity. Utilizing 20 μm height DEP chambers, “almost complete” and …

Selection For Non-Amyloidogenic Mutants Of Islet Amyloid Polypeptide (Iapp) Identifies An Extended Region For Amyloidogenicity, Ayano Fox, Thibaut Snollaerts, Camille Errecart Casanova, Anastasia Calciano, Luiza A. Nogaj, David A. Moffet

Chemistry and Biochemistry Faculty Works

The aggregation of the 37-residue polypeptide IAPP, as either insoluble amyloid or as small oligomers, appears to play a direct role in the death of pancreatic β-islet cells in type II diabetes. While IAPP has been known to be the primary component of type II diabetes amyloid, the molecular interactions responsible for this aggregation have not been identified. To identify the aggregation-prone region(s), we constructed a library of randomly generated point mutants of IAPP. This mutant IAPP library was expressed in E. coli as genetic fusions to the reporter protein enhanced green fluorescent protein (EGFP). Because IAPP aggregates rapidly, both …