Biochemistry, Biophysics, and Structural Biology Commons^™

Calcium Transport Inhibition, Stimulation, And Light Dependent Modulation Of The Skeletal Calcium Release Channel (Ryr1) By The Prototropic Forms Of Pelargonidin, Thomas Joseph Dornan

Dissertations and Theses

The principle calcium regulator in the muscle cell is the calcium ion release channel (RyR). Improper calcium homeostasis in the muscle cell is the foundation of many pathological states and has been targeted as a contributing factor to ventricular tachycardia, which is known to precede sudden cardiac arrest.

Numerous endogenous and exogenous compounds can affect the way RyR regulates calcium. In this study the anthocyanidin Pelargonidin (Pg), an important natural colorant and dietary antioxidant, is evaluated for its effect on regulating the transport of calcium through the RyR1 of skeletal muscle sarcoplasmic reticulum. Pelargonidin undergoes time dependent structural changes in …

High-Performance Polymer Monoliths For Capillary Liquid Chromatography, Pankaj Aggarwal

Theses and Dissertations

This dissertation focuses on improving the chromatographic efficiency of polymeric organic monoliths by characterizing and optimizing the bed morphology. In-situ characterization techniques such as capillary flow porometry (CFP), 3-dimensional scanning electron microscopy (3D SEM) and conductivity measurements were developed and implemented to quantitatively characterize the morphology of poly(ethylene glycol) diacrylate (PEGDA) monoliths. The CFP measurements for monoliths prepared by the same procedure in capillaries with different diameters (i.e., 75, 150, and 250 μm) clearly showed a change in average through-pore size with capillary diameter, thus, certifying the need for in-situ measurement techniques. Serial sectioning and imaging of PEGDA monoliths using …

Development And In Silico Evaluation Of Large-Scale Metabolite Identification Methods Using Functional Group Detection For Metabolomics, Joshua M. Mitchell, Teresa W-M Fan, Andrew N. Lane, Hunter N. B. Moseley

Molecular and Cellular Biochemistry Faculty Publications

Large-scale identification of metabolites is key to elucidating and modeling metabolism at the systems level. Advances in metabolomics technologies, particularly ultra-high resolution mass spectrometry (MS) enable comprehensive and rapid analysis of metabolites. However, a significant barrier to meaningful data interpretation is the identification of a wide range of metabolites including unknowns and the determination of their role(s) in various metabolic networks. Chemoselective (CS) probes to tag metabolite functional groups combined with high mass accuracy provide additional structural constraints for metabolite identification and quantification. We have developed a novel algorithm, Chemically Aware Substructure Search (CASS) that efficiently detects functional groups within …

Some Effects Of The Human Genome Project On The Erdős Collaboration Graph, Chris Fields

Journal of Humanistic Mathematics

The Human Genome Project introduced large-scale collaborations involving dozens to hundreds of scientists into biology. It also created a pressing need to solve discrete mathematics problems involving tens of thousands of elements. In this paper, we use minimal path lengths in the Erdős Collaboration Graph between prominent individual researchers as a measure of the distance between disciplines, and we show that the Human Genome Project brought laboratory biology as a whole closer to mathematics. We also define a novel graph reduction method and a metric that emphasizes the robustness of collaborative connections between researchers; these can facilitate the analysis of …

Water Use And Water Tolerance In The Riparian Species American Sycamore (Plantanus Occidentalis L.) And How Its Physiology Compares To The Coexisting Black Willow (Salix Nigra Marshall), Eric Duncan

Dissertations, Theses and Capstone Projects

This study set out to compare aspects of water use and tolerance to inundation and drought of two riparian tree species, Platanus occidentalis L. and Salix nigra Marshall. The main goal of the study was to better understand the two species and assess the suitability of P. occidentalis for use in riparian restoration. To accomplish this, potted individuals were subjected to treatments of drought, inundation, or control, under greenhouse conditions, and separately, in order to estimate water use in trees, season-long diurnal measurements of sap flux (using Granier type thermal dissipation probes) were determined in situ for trees growing in …

Metagenomic Identification Of A Novel Salt Tolerance Gene From The Human Gut Microbiome Which Encodes A Membrane Protein With Homology To A Brp/Blh-Family Beta-Carotene 15,15'-Monooxygenase, Eamonn P. Culligan, Roy D. Sleator, Julian R. Marchesi, Colin Hill

Department of Biological Sciences Publications

The human gut microbiome consists of at least 3 million non-redundant genes, 150 times that of the core human genome. Herein, we report the identification and characterisation of a novel stress tolerance gene from the human gut metagenome. The locus, assigned brpA, encodes a membrane protein with homology to a brp/blh-family β-carotene monooxygenase. Cloning and heterologous expression of brpA in Escherichia coli confers a significant salt tolerance phenotype. Furthermore, when cultured in the presence of exogenous β-carotene, cell pellets adopt a red/orange pigmentation indicating the incorporation of carotenoids in the cell membrane.

Megadalton-Node Assembly By Binding Of Skb1 To The Membrane Anchor Slf1, Lin L. Deng, Ruth Kabeche, Ning Wang, Jian-Qiu Wu, James B. Moseley

Dartmouth Scholarship

The plasma membrane contains both dynamic and static microdomains. Given the growing appreciation of cortical microdomains in cell biology, it is important to determine the organizational principles that underlie assembly of compartmentalized structures at the plasma membrane. The fission yeast plasma membrane is highly compartmentalized by distinct sets of cortical nodes, which control signaling for cell cycle progression and cytokinesis. The mitotic inhibitor Skb1 localizes to a set of cortical nodes that provide spatial control over signaling for entry into mitosis. However, it has been unclear whether these nodes contain other proteins and how they might be organized and tethered …

[Accepted Article Manuscript Version (Postprint)] Auxin Input Pathway Disruptions Are Mitigated By Changes In Auxin Biosynthetic Gene Expression In Arabidopsis, Gretchen Spiess, Amanda Hausman, Peng Yu, Jerry Cohen, Rebekah Rampey, Bethany Zolman

Biology Department Faculty Works

Auxin is a phytohormone involved in cell elongation and division. Levels of indole-3-acetic acid (IAA), the primary auxin, are tightly regulated through biosynthesis, degradation, sequestration, and transport. IAA is sequestered in reversible processes by adding amino acids, polyol or simple alcohols, or sugars, forming IAA conjugates, or through a two-carbon elongation forming indole-3-butyric acid. These sequestered forms of IAA alter hormone activity. To gain a better understanding of how auxin homeostasis is maintained, we have generated Arabidopsis (Arabidopsis thaliana) mutants that combine disruptions in the pathways, converting IAA conjugates and indole-3-butyric acid to free IAA. These mutants show phenotypes indicative …

Megatevs: Single-Chain Dual Nucleases For Efficient Gene Disruption, Jason M Wolfs, Matthew Dasilva, Sarah E Meister, Xu Wang, Caroline Schild-Poulter, David R. Edgell

Biochemistry Publications

Targeting gene disruptions in complex genomes relies on imprecise repair by the non-homologous end-joining DNA pathway, creating mutagenic insertions or deletions (indels) at the break point. DNA end-processing enzymes are often co-expressed with genome-editing nucleases to enhance the frequency of indels, as the compatible cohesive ends generated by the nucleases can be precisely repaired, leading to a cycle of cleavage and non-mutagenic repair. Here, we present an alternative strategy to bias repair toward gene disruption by fusing two different nuclease active sites from I-TevI (a GIY-YIG enzyme) and I-OnuI E2 (an engineered meganuclease) into a single polypeptide chain. In vitro, …

Evaluation Of Eight Live Attenuated Vaccine Candidates For Protection Against Challenge With Virulent Mycobacterium Avium Subspecies Paratuberclosis In Mice, John Bannantine, Jamie L. Everman, Sasha J. J.Rose, Lmar Babrak, Robab Katani, Raul G. Barletta, Adel M. Talaat, Yrjö J. Gröhn, Yung-Fu Chang, Vivek Kapur, Luiz E. Bermudez

School of Veterinary and Biomedical Sciences: Faculty Publications

Johne's disease is caused by Mycobacterium avium subsp. paratuberculosis (MAP), which results in serious economic losses worldwide in farmed livestock such as cattle, sheep, and goats. To control this disease, an effective vaccine with minimal adverse effects is needed. In order to identify a live vaccine for Johne's disease, we evaluated eight attenuated mutant strains of MAP using a C57BL/6 mouse model. The persistaence of the vaccine candidates was measured at 6, 12, and 18 weeks post vaccination. Only strains 320, 321, and 329 colonized both the liver and spleens up until the 12-week time point. The remaining five mutants …

Density Functional Theory Modeling Of It-Stacking And Electrophilic Donor-Acceptor Interactions With Application To Therapeutic Targeting Of Zinc-Finger Proteins, Patricia Beall Lutz

Chemistry & Biochemistry Theses & Dissertations

Several viruses, including viruses that cause cancer, contain conserved zinc finger (ZF) proteins that are essential for viral reproduction, making them attractive drug targets for cancer and viral treatment. ZFs are small protein domains that have Zn²⁺ tetrahedrally coordinated to at least 2 Cys and His. They form three classes of ZFs depending on the amino acid ligands, CCHH, CCCH, and CCCC. Zn²⁺ is stable towards redox reactions; however, the Cys thiolates are redox active. Oxidation of the Cys thiolates release Zn²⁺, and the ZF loses its tertiary structure and can no longer bind DNA or …

Characterization Of Lipid-Anchored Inhibitor Rulers As A Measure Of Enzyme Topography In Coagulation Enzyme Factor Xa, Robin C. Conley

PCOM Biomedical Studies Student Scholarship

The blood coagulation cascade is activated under various circumstances such as an injury. This system involves a tightly regulated series of events. The enzymes involved assemble with their respective cofactors on lipid membranes to reach their full procoagulant complex potential. The coagulation cascade is divided into intrinsic and extrinsic portions, both of which converge into a common pathway with the activation of factor X (FX_a). FX_a is a very important part of the blood coagulation cascade because its activation is primarily responsible for thrombin generation. Activation of FX_a alone is, however, insufficient to produce a fully …

Using The Microsoft Kinect To Assess Human Bimanual Coordination, Joshua James Liddy

Open Access Theses

Optical marker-based systems are the gold-standard for capturing three-dimensional (3D) human kinematics. However, these systems have various drawbacks including time consuming marker placement, soft tissue movement artifact, and are prohibitively expensive and non-portable. The Microsoft Kinect is an inexpensive, portable, depth camera that can be used to capture 3D human movement kinematics. Numerous investigations have assessed the Kinect's ability to capture postural control and gait, but to date, no study has evaluated it's capabilities for measuring spatiotemporal coordination. In order to investigate human coordination and coordination stability with the Kinect, a well-studied bimanual coordination paradigm (Kelso, 1984, Kelso; Scholz, & …

Investigation Into The Control Of Melittin Secondary Structure And Antimicrobial Activity, Zachary B. Molinets

Open Access Theses

Antimicrobial resistance has been an exponentially growing problem since the discovery of antibiotics. Antibiotics have been misused for many years and this misuse has grown into a real problem for the medical community. While there are countless safeguards to prevent infection by a resistant strain of bacteria, there are still many plagued by it and must be treated with sometimes dangerous antibiotics. Melittin, along with many other peptides, contain potent antimicrobial properties, but are also toxic toward enthrocytes. The control of the secondary structure of peptides provides the key to adjusting their activity.

Key Residues Of Human Cytoplasmic Protein Tyrosine Phosphatase-A And -B For Substrate Binding And Specificity, Byunghyun Park

Open Access Theses

Reversible tyrosine phosphorylation plays an important role in signaling pathways that are essential for regulating cellular growth, differentiation and metabolism. Moreover, several human diseases such as diabetes, obesity and cancers are associated with the deregulation of protein tyrosine phosphatases (PTPs). Several studies provide evidence that PTPs not only contribute to cellular differentiation, but over-expression of these molecules also leads to transformation of non-transfomed cells as well. Based on these results, designing specific PTP inhibitors may ultimately function as potential therapeutic agents to treat various diseases including cancer, diabetes, and autoimmune diseases. EphA2 is a receptor tyrosine kinase which is hypo-phosphorylated …

Abcb11 Functions With B1 And B19 To Regulate Rootward Auxin Transport, Jesica Elyse Reemmer

Open Access Theses

Auxin transport is essential for the architecture and development of erect plants. In a network of transporters directing auxin flows, ATP-Binding Cassette (ABC) transporters are a ubiquitous family of proteins that actively transport important substrates, including auxins, across the plasma membrane. ABCB1 and ABCB19 have been shown to account for the majority of rootward auxin transport, but residual fluxes to the root tip in Arabidopsis b1b19 double mutants implies the involvement of at least one additional auxin transporter in this process. Of specific interest, the severe dwarfism seen in abcb1abcb19 is strikingly reminiscent of that seen in mutants defective in …

Efficacy Of Cleaning Method For Removal Of Exogenous Welding Fume Contamination From Nail Tissue Prior To Use As A Biomarker For Welding Fume Manganese Exposure, Jeffrey Corral Bainter

Open Access Theses

Nail tissue has been proposed as a biomarker for body burden of occupational exposure to manganese from welding fumes. Though recent studies have shown correlation between manganese exposure and both nail tissue concentration as well as concentrations in dopaminergic regions of the brain, concerns of the validity of nail tissue as a biomarker have arisen due to the potential for exogenous contamination of Mn to undermine the quantization of endogenous Mn in nail. Previous studies have used a cleaning method of 1% Triton X-100 surfactant plus sonication in order to attempt to remove exogenous welding fume contamination. Determination of the …

1,25-Dihydroxyvitamin D Alters Lipid Metabolism And Epithelialto- Mesenchymal Transition In Metastatic Epithelial Breast Cancer Cells, Alle Nicole Barnard

Open Access Theses

Evidence suggests that high vitamin D status (marked by serum 25-hydroxyvitamin D, 25(OH)2 D) is associated with a decreased risk of breast cancer. It has been established that 1,25-dihydroxyvitamin D (1,25(OH) 2 D) can alter glycolysis and the Krebs cycle of breast cancer cells (Jiang et al., 2010; Zheng et al., 2013) but little information is available on 1,25(OH) 2 D's alterations of lipid metabolism in breast cancer cells. Thus, the current research investigates if there was an effect of 1,25(OH) 2 D on proteins that regulate lipid metabolism in MCF10A, MCF10A-ras, MCF10CA1h, and MCF10CA1a epithelial breast cancer cells. While …

Synthesis And Dna-Binding Studies With Two Stericallyfriendly Porphyrin Frameworks, Srijana Ghimire

Open Access Dissertations

Peripheral substituents on cationic porphyrins play a significant role during binding with DNA hosts. Possible applications of these systems in photodynamic therapy as well as in anti-bacterial and anti-cancer therapies motivate the binding studies. For characterizing DNA binding motifs different methods are useful including absorption, emission, and circular dichroism spectroscopies, as well as viscometry and X-ray crystallography. With the classic H₂ T₄ porphyrin, or 5,10,15,20-tetra(N-methylpyridinium-4-yl)porphyrin, the mode of binding varies with the base composition of the DNA host. The porphyrin binds adenine-thymine rich sequences externally whereas intercalation occurs in guanine-cytosine rich sequences. The McMillin group has made some …

Gmos: Are They A Regulatory Or Food Safety Issue?, Richard E. Goodman

Department of Food Science and Technology: Faculty Publications

Most of the genetic diversity that has improved agricultural production throughout the history of farming was developed through natural mutations and selective breeding. However, since the early 1900s plant scientists have used chemical and radiation mutagenesis to increase genetic diversity (18). We know that the majority of mutations are harmful, and plant breeders work hard to select only those that are beneficial. This process has helped feed a growing human population, which is estimated to have been 300 million 2,000 years ago and is now more than 7 billion. However, methods used in the past to improve agricultural production are …

Bioinformatic Solutions To Complex Problems In Mass Spectrometry Based Analysis Of Biomolecules, Ryan M. Taylor

Theses and Dissertations

Biological research has benefitted greatly from the advent of omic methods. For many biomolecules, mass spectrometry (MS) methods are most widely employed due to the sensitivity which allows low quantities of sample and the speed which allows analysis of complex samples. Improvements in instrument and sample preparation techniques create opportunities for large scale experimentation. The complexity and volume of data produced by modern MS-omic instrumentation challenges biological interpretation, while the complexity of the instrumentation, sample noise, and complexity of data analysis present difficulties in maintaining and ensuring data quality, validity, and relevance. We present a corpus of tools which improves …

Experimental And Computational Analysis Of The Synucleins, Agatha Munyanyi

Theses and Dissertations in Biomedical Sciences

The synuclein proteins α, β and γ which are located in the brain, have been a subject of intense research. Of particular interest is α-synuclein, which is found in misfolded forms in Lewy bodies that are associated with Parkinson's disease. Despite the efforts of researchers across the world, the physiological structure and function of the synucleins remains elusive. In recent years, highly controversial reports by some investigators indicate that in its natural form, α-synuclein exists as a tetramer instead of as an intrinsically unstructured monomer. This dissertation presents results of the experimental and computational analysis of the synucleins. First, we …

The Interrelationship Of Brca1 185delag, Interleukin-1Β, And Ovarian Oncogenesis, Kamisha Woolery

USF Tampa Graduate Theses and Dissertations

While the etiology of ovarian cancer (OC) is not completely understood, evidence suggests that chronic inflammation may promote malignant transformation. However, familial history remains the strongest risk factor for developing OC and is associated with germline BRCA1 mutations, such as the 185delAG mutation. Normal human ovarian surface epithelial cells expressing the 185delAG mutant, BRAT, exhibit molecular and pathological changes that may contribute to OC oncogenesis. In the current study, I sought to determine whether BRAT could promote an inflammatory phenotype by investigating BRAT's impact on the expression of the proinflammatory cytokine, Interleukin-1β (IL-1β). Using a culture model system of normal …

Rna Detection Technology For Applications In Marine Science: Microbes To Fish, Robert Michael Ulrich

USF Tampa Graduate Theses and Dissertations

The accurate identification of taxa from mixed assemblages using genetic analysis remains an important field of molecular biology research. The common principle behind the development of numerous documented genetic detection technologies is to exploit specific nucleotide sequences inherent to each taxon. This body of work focuses on practical applications of real-time nucleic acid sequence-based amplification (RT-NASBA) in marine science, and is presented in four case studies. Each study represents novel work in the genetic identification of respective taxa of interest using RT-NASBA. Two case studies documented the development of an assay targeting mitochondrial 16S rRNA to discern legally salable grouper …

Laser-Induced Fluorescence Imaging Of Calcium And Barium Ion Beams In The Second Vacuum Stage Of A Commercial Inductively Coupled Plasma Mass Spectrometer, Alisa Jane Edmund

Theses and Dissertations

Inductively coupled plasma-mass spectrometers (ICP-MS) have become the workhorses of many analytical labs over the past few decades. Despite the instruments' high sensitivities and low detection limits there is still a demand for improvements in several aspects of their performance. One area of improvement is in the understanding of "space charge effects" Space charge effects are classified as problems associated with the ion beam. Problems are created when the mutual repulsions of the ions make consistent focusing of the ion beam difficult. This is particularly problematic with samples containing a low concentration analyte contained within a high salt solution matrix, …

Synthesis, Characterization And Mechanistic Studies Of Biomolecules@Mesomofs, Yao Chen

USF Tampa Graduate Theses and Dissertations

Encapsulation of biomolecules is of great interest to research advances related to biology, physiology, immunology, and biochemistry, as well as industrial and biomedical applications such as drug delivery, biocatalysis, biofuel, food and cosmetics. Encapsulation provides functional characteristics that are not fulfilled by free biomolecules and stabilizes the fragile biomolecules. In terms of biocatalysis, solid support can often enhance the stability of enzymes, as well as facilitate separation and recovery for reuse while maintaining activity and selectivity. Various kinds of materials have been used for encapsulation of biomolecules, among which, porous materials are an important group. Metal-organic frameworks (MOFs) have attracted …

Identification Of Cd24 As A Cancer Stem Cell Marker In Human Nasopharyngeal Carcinoma, Chun-Hung Yang, Hui-Ling Wang, Yi-Sheng Lin, K. P. Shravan Kumar, Hung-Chi Lin, Chih-Jung Chang, Chia-Chen Lu, Tsung-Teng Huang, Jan Martel, David M. Ojcius, Yu-Sun Chang, John Ding-E. Young, Hsin-Chih Lai

All Dugoni School of Dentistry Faculty Articles

Cancer stem cells (CSCs) represent a unique sub-population of tumor cells with the ability to initiate tumor growth and sustain self-renewal. Although CSC biomarkers have been described for various tumors, only a few markers have been identified for nasopharyngeal carcinoma (NPC). In this study, we show that CD24⁺ cells isolated from human NPC cell lines express stem cell genes (Sox2, Oct4, Nanog, Bmi-1, and Rex-1), and show activation of the Wnt/β-catenin signaling pathway. CD24⁺ cells possess typical CSC characteristics that include enhanced cell proliferation, increased colony and sphere formation, maintenance of cell differentiation potential in prolonged …

Anthropogenic Litter In Urban Freshwater Ecosystems: Distribution And Microbial Interactions, Timothy Hoellein, Miguel Rojas, Adam Pink, Joseph Gasior, John J. Kelly

Biology: Faculty Publications and Other Works

Accumulation of anthropogenic litter (i.e. garbage; AL) and its ecosystem effects in marine environments are well documented. Rivers receive AL from terrestrial habitats and represent a major source of AL to marine environments, but AL is rarely studied within freshwater ecosystems. Our objectives were to 1) quantify AL density in urban freshwaters, 2) compare AL abundance among freshwater, terrestrial, and marine ecosystems, and 3) characterize the activity and composition of AL biofilms in freshwater habitats. We quantified AL from the Chicago River and Chicago's Lake Michigan shoreline, and found that AL abundance in Chicago freshwater ecosystems was comparable to previously …

Membrane Inlet Mass Spectrometry Reveals That Ceriporiopsis Subvermispora Bicupin Oxalate Oxidase Is Inhibited By Nitric Oxide, Ellen W. Moomaw, Richard Uberto, Tu Chingkuang

Faculty Articles

Membrane inlet mass spectrometry (MIMS) uses a semipermeable membrane as an inlet to a mass spectrometer for the measurement of the concentration of small uncharged molecules in solution. We report the use of MIMS to characterize the catalytic properties of oxalate oxidase (E.C. 1.2.3.4) from Ceriporiopsis subvermispora (CsOxOx). Oxalate oxidase is a manganese dependent enzyme that catalyzes the oxygen-dependent oxidation of oxalate to carbon dioxide in a reaction that is coupled with the formation of hydrogen peroxide. CsOxOx is the first bicupin enzyme identified that catalyzes this reaction. The MIMS method of measuring OxOx activity involves continuous, real-time direct detection …

Novel Cell Cycle Proteins In Apicomplexan Parasites, Carrie Butler

USF Tampa Graduate Theses and Dissertations

Apicomplexans are responsible for major human diseases such as toxoplasmosis caused by Toxoplasma gondii (T. gondii) and the deadliest form of malaria caused by Plasmodium falciparum (P. falciparum). The genomes of these pathogens are now sequenced ushering in a new era of drug development. A major hurdle to exploiting this genome resource is that a large number of the encoded genes are "hypotheticals" and have yet to be characterized. Hypothetical proteins comprise roughly half of the predicted gene complement of T. gondii and P. falciparum and represent the largest class of uniquely functioning proteins in these parasites.

Following the idea …