Molecular Biology Commons^™

Synthesis Of Rhamnosylated Arginine Glycopeptides And Determination Of The Glycosidic Linkage In Bacterial Elongation Factor P, Siyao Wang, Leo Corcilius, Phillip B. Sharp, Andrei Rajkovic, Michael Ibba, Benjamin L. Parker, Richard J. Payne

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

A new class of N-linked protein glycosylation – arginine rhamnosylation – has recently been discovered as a critical modification for the function of bacterial elongation factor P (EF-P). Herein, we describe the synthesis of suitably protected α- and β-rhamnosylated arginine amino acid “cassettes” that can be directly installed into rhamnosylated peptides. Preparation of a proteolytic fragment of Pseudomonas aeruginosa EF-P bearing both α- and β-rhamnosylated arginine enabled the unequivocal determination of the native glycosidic linkage to be α through 2D NMR and nano-UHPLC-tandem mass spectrometry studies.

Inflammatory Gene Expression In Women Diagnosed With Polycystic Ovarian Syndrome, Myles K. Taylor

Electronic Theses & Dissertations

Polycystic ovarian syndrome (PCOS) is a common endocrine disorder in women of reproductive age. The pathophysiology of PCOS has conventionally thought to originate from androgen excess. However, recent evidence suggests that androgen excess is a downstream consequence to inflammatory dysregulation and subsequent metabolic abnormalities. Inflammatory mRNA gene expression of TNFa and IL-1ß in mononuclear cells isolated from women diagnosed with PCOS was explored using qPCR. Additionally, the correlations between body mass index (BMI) and fasting glucose on mRNA expression of TNFa and IL-1ß were explored. mRNA expression of both TNFa and IL-1ß were found to be significantly higher in PCOS …

Characterization Of Vesicular Monoamine Transporter 2 And Its Role In Parkinson's Disease Pathogenesis Using Drosophila, Antonio Joel Tito Jr., Sheng Zhang

Dissertations & Theses (Open Access)

Parkinson’s disease (PD) is a progressive neurodegenerative disorder caused by the selective loss of the dopaminergic neurons in the Substantia nigra pars compacta region of the brain. PD is also the most common neurodegenerative disorder and the second most common movement disorder. PD patients exhibit the cardinal symptoms, including tremor of the extremities, rigidity, slowness of movement, and postural instability, after 70-80% of DA neurons degenerate. It is, therefore, imperative to elucidate the underlying mechanisms involved in the selective degeneration of DA neurons. Although increasing numbers of PD genes have been identified, why these largely widely expressed genes induce …

Consensus Paper: Cerebellar Development., K Leto, M Arancillo, Ebe Becker, A Chiang, Et Al.

Faculty Publications

The development of the mammalian cerebellum is orchestrated by both cell-autonomous programs and inductive environmental influences. Here, we describe the main processes of cerebellar ontogenesis, highlighting the neurogenic strategies used by developing progenitors, the genetic programs involved in cell fate specification, the progressive changes of structural organization, and some of the better-known abnormalities associated with developmental disorders of the cerebellum.

The Complex Evolutionary History Of Aminoacyl-Trna Synthetases, Anargyros Chaliotis, Panayotis Vlastaridis, Dimitris Mossialos, Michael Ibba, Hubert D. Becker, Constantinos Stathopoulos, Grigorios D. Amoutzias

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Aminoacyl-tRNA synthetases (AARSs) are a superfamily of enzymes responsible for the faithful translation of the genetic code and have lately become a prominent target for synthetic biologists. Our large-scale analysis of >2500 prokaryotic genomes reveals the complex evolutionary history of these enzymes and their paralogs, in which horizontal gene transfer played an important role. These results show that a widespread belief in the evolutionary stability of this superfamily is misconceived. Although AlaRS, GlyRS, LeuRS, IleRS, ValRS are the most stable members of the family, GluRS, LysRS and CysRS often have paralogs, whereas AsnRS, GlnRS, PylRS and SepRS are often absent …

Gene Expression Profiling And The Role Of Hsf1 In Ovarian Cancer In 3d Spheroid Models, Trillitye Paullin

USF Tampa Graduate Theses and Dissertations

Ovarian cancer is the most lethal gynecological cancer, with over 200,000 women diagnosed each year and over half of those cases leading to death. These poor statistics are related to a lack of early symptoms and inadequate screening techniques. This results in the cancer going undetected until later stages when the tumor has metastasized through a process that requires the epithelial to mesenchymal transition (EMT). In lieu of traditional monolayer cell culture, EMT and cancer progression in general is best characterized through the use of 3D spheroid models. In this study, we examine gene expression changes through microarray analysis in …

The Rodent Parvovirus H-1 And Its Potential In Combination With Tien Hsien Liquid As A Treatment For Breast Cancer, Hayley I. Spires

Selected Honors Theses

Breast cancer is responsible for 12% of cancer diagnoses each year, and it is the leading cause of cancer-related mortality in women worldwide. Current treatments have provided some success in combatting the disease but are not considered a final solution. The framework for how researchers approach cancer has continued to change, and this includes the introduction of oncolytic viruses as novel therapeutics for cancer. The rodent parvovirus H-1 has shown strong potential in clinical and subclinical trials, but its S phase dependency limits its usefulness against cancer stem cell populations. Tien Hsien Liquid is commerically available, nontoxic, and has shown …

Understanding The Differences Between Neuronal Calcium Sensor Proteins: A Comparison Of Neurocalcin Delta And Hippocalcin, Jeffrey M. Viviano

Graduate School of Biomedical Sciences Theses and Dissertations

Many neuronal functions, including learning and memory are driven by changes in intracellular Ca2+concentrations. The Neuronal Calcium Sensor (NCS) family of proteins is responsible for mediating the response to calcium. They are typically comprised of 4 EF hands; of which EF 2, 3, and 4 bind calcium.

Hypothesis: NCS proteins carry out unique, non-overlapping functions, and that specific characteristics of the family can be mapped to precise regions of the proteins.

Results: The effect on the following properties were investigated primarily on two highly similar NCS proteins, Neurocalcin Delta (NCALD) and Hippocalcin (HPCA): (1) Response to calcium was determined through …

Contribution Of Epithelial Hypoxia Signaling To Pulmonary Fibrosis: Role Of Fak1 And Galectin-1 As Driver Molecules, Jaymin J. Kathiriya

USF Tampa Graduate Theses and Dissertations

Idiopathic Pulmonary Fibrosis (IPF) is a deadly disease of unknown origin, which causes 80,000 deaths every year in the US and Europe combined. Unknown etiology and late diagnosis, combined with limited treatment options, contribute to a dismal survival rate of 3-5 years post diagnosis. Although molecular mechanisms underlying IPF pathogenesis and progression have been studied for over two decades, lack of in vivo models that recapitulate chronic, progressive, and irreversible nature of IPF have contributed to limited therapeutic success in clinical trials. Currently, only two drugs, Pirfenidone and Nintedanib, are approved for IPF treatment in the US, with their efficacy …

An Arginine Finger Regulates The Sequential Action Of Asymmetrical Hexameric Atpase In The Double-Stranded Dna Translocation Motor, Zhengyi Zhao, Gian Marco De-Donatis, Chad T. Schwartz, Huaming Fang, Jingyuan Li, Peixuan Guo

Pharmaceutical Sciences Faculty Publications

Biological motors are ubiquitous in living systems. Currently, how the motor components coordinate the unidirectional motion is elusive in most cases. Here, we report that the sequential action of the ATPase ring in the DNA packaging motor of bacteriophage ϕ29 is regulated by an arginine finger that extends from one ATPase subunit to the adjacent unit to promote noncovalent dimer formation. Mutation of the arginine finger resulted in the interruption of ATPase oligomerization, ATP binding/hydrolysis, and DNA translocation. Dimer formation reappeared when arginine mutants were mixed with other ATPase subunits that can offer the arginine to promote their interaction. Ultracentrifugation …

Identification And Characterization Of The Anticancer Potential Of Indigenous Medicinal Plants Of The Arabian Peninsula, Sameera Omar Mohammed Saeed Balhamar

Theses

Indigenous plant species historically used for their medicinal properties are a tremendous source for bringing newer and safer drugs to the market. A concerted effort is needed to characterize their medicinal potential and identify new molecules that could be exploited in modern medicine. The current study was undertaken to study the anticancer properties of several indigenous plants that are used by the local population of the Arabian Peninsula and beyond for various medicinal purposes. Towards this end, we acquired different plant extracts from five plants, namely Boswellia sacra (BS), Cleome droserifolia (CD), Teucrium muscatensis (TM), Orchadenus arabicus (OA), and Acredocarpus …

Presence Of A Prophage Determines Temperature-Dependent Capsule Production In Streptococcus Pyogenes., Leslie Brown, Jeong-Ho Kim, Kyu Hong Cho

Biochemistry and Molecular Medicine Faculty Publications

A hyaluronic acid capsule is a major virulence factor in the pathogenesis of Streptococcus pyogenes. It acts as an anti-phagocytic agent and adhesin to keratinocytes. The expression of the capsule is primarily regulated at the transcriptional level by the two-component regulatory system CovRS, in which CovR acts as a transcriptional repressor. The covRS genes are frequently mutated in many invasive strains, and a subset of the invasive CovRS mutants does not produce a detectable level of the capsule at 37 °C, but produces a significant amount of the capsule at sub-body temperatures. Here, we report that a prophage has a …

Isoacceptor Specific Characterization Of Trna Aminoacylation And Misacylation In Vivo, Kyle Mohler, Rebecca Mann, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Amino acid misincorporation during protein synthesis occurs due to misacylation of tRNAs or defects in decoding at the ribosome. While misincorporation of amino acids has been observed in a variety of contexts, less work has been done to directly assess the extent to which specific tRNAs are misacylated in vivo, and the identity of the misacylated amino acid moiety. Here we describe tRNA isoacceptor specific aminoacylation profiling (ISAP), a method to identify and quantify the amino acids attached to a tRNA species in vivo. ISAP allows compilation of aminoacylation profiles for specific isoacceptors tRNAs. To demonstrate the efficacy and …

Maintenance Of Transcription-Translation Coupling By Elongation Factor P, Sara Elgamal, Irina Artsimovitch, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Under conditions of tight coupling between translation and transcription, the ribosome enables synthesis of full-length mRNAs by preventing both formation of intrinsic terminator hairpins and loading of the transcription termination factor Rho. While previous studies have focused on transcription factors, we investigated the role of Escherichia coli elongation factor P (EF-P), an elongation factor required for efficient translation of mRNAs containing consecutive proline codons, in maintaining coupled translation and transcription. In the absence of EF-P, the presence of Rho utilization (rut) sites led to an ~30-fold decrease in translation of polyproline-encoding mRNAs. Coexpression of the Rho inhibitor Psu …

Insights Into The Molecular Roles Of Heparan Sulfate Proteoglycans (Hspgs-Syndecans) In Autocrine And Paracrine Growth Factor Signaling In The Pathogenesis Of Hodgkin's Lymphoma, Rajendra Ghardbaran

Publications and Research

Syndecans (SDC, SYND) comprise a group of four structurally related type 1 transmembrane heparan sulfate proteoglycans (HSPGs) that play important roles in tumorigenic processes. SDCs exert signaling via their protein cores and their conserved transmembrane and cytoplasmic domains or by forming complexes with growth factors (GFs). In classical Hodgkin's lymphoma (cHL), a lymphoid neoplasm of predominantly B cell origin, SDC1 and SDC4 are the active SDCs, and a number of GF (vascular endothelial growth factor, fibroblast growth factor, etc.) signaling pathways have been studied. However, despite extensive pre-clinical and clinical research on SDC-mediated GF signaling in many cancer types, there …

Regulation Of Nachrs And Stemness By Nicotine And E-Cigarettes In Nsclc, Courtney Schaal

USF Tampa Graduate Theses and Dissertations

Lung cancer is the leading cause of cancer-related death in both men and women, nationally and internationally and kills more people each year than breast, prostate, and colon cancers combined. Non-small cell lung carcinoma (NSCLC) is the most common histological subtype of lung cancer, and accounts for 85% of all cases. Cigarette smoking is the single greatest risk factor for lung cancer, and is correlated with 80-90% of all lung cancer deaths. Nicotine, the addictive component of tobacco smoke, is not a carcinogen and cannot initiate tumors itself; however, it is known to act as a tumor promoter, by enhancing …

Dynamic Host-Pathogen Interactions Result In Fungal Epitope Unmasking, Alex Hopke

Electronic Theses and Dissertations

Molecular camouflage is used by a diverse set of pathogens to disguise their identity and avoid recognition by protective host receptors. The opportunistic fungal pathogen Candida albicans is a good example, as it masks the inflammatory component β-glucan in its cell wall to evade detection by the immune receptor Dectin-1. Interestingly, it has been seen that β-glucan becomes unmasked during infection in vivo, though the underlying mechanisms remained unclear. Exposure levels of this epitope may be important, as Dectin-1 mediates protection from some strains of C. albicans and alterations in the organization and composition of the Candida cell wall …

Cal And Magi Pdz Protein Regulation Of Crfr1 And 5-Ht2ar Trafficking And Signaling, Maha Mahmoud Hammad

Electronic Thesis and Dissertation Repository

PDZ (PSD95/Disc Large/Zona Occludens) domain-containing proteins are scaffolding proteins that play important roles in regulating the activity of G protein-coupled receptors. Corticotropin Releasing Factor Receptor 1 (CRFR1) and Serotonin 2A Receptor (5-HT_2AR) are two GPCRs that are commonly associated with mental disorders. Both receptors also contain a class I PDZ-binding motif at the carboxyl terminal tail. In the first chapter, we investigate the effects of CAL (CFTR-associated ligand) on regulating the trafficking and signaling of CRFR1. We demonstrate a role for CAL in inhibiting CRFR1 endocytosis, cell surface expression, and CRF-mediated ERK1/2 signaling via the CRFR1 PDZ-binding motif. …

Design And Development Of A Plasmid Vector For Protein Expression And Purification, Mahima Grover, Craig Sweet, David H. Thompson

The Summer Undergraduate Research Fellowship (SURF) Symposium

Production and isolation of proteins are difficult, costly and time-consuming processes. The aim of this project is for the development of plasmids, which allow for streamlined production and isolation of proteins. To allow for modular insertion of varying segments of DNA we are using ‘recursive directional ligation by plasmid reconstruction’. This technique uses type II restriction endonucleases, which cut downstream from their recognition site allowing multiple insertions without losing a restriction site. Using this process, we can ligate multiple DNA sequences together and express them to be able to construct a scar less fusion protein. In order to accomplish this, …

Structural And Molecular Analysis Of A Protective Epitope Of Lyme Disease Antigen Ospa And Antibody Interactions, Shivender Shandilya, Nese Kurt Yilmaz, Ejemel Monir, Andrew Sadowski, William D. Thomas, Mark S. Klempner, Celia A. Schiffer, Yan Wang

Celia A. Schiffer

The murine monoclonal antibody LA-2 recognizes a clinically protective epitope on outer surface protein (OspA) of Borrelia burgdorferi, the causative agent of Lyme disease in North America. Human antibody equivalence to LA-2 is the best serologic correlate of protective antibody responses following OspA vaccination. Understanding the structural and functional basis of the LA-2 protective epitope is important for developing OspA-based vaccines and discovering prophylactic antibodies against Lyme disease. Here, we present a detailed structure-based analysis of the LA-2/OspA interaction interface and identification of residues mediating antibody recognition. Mutations were introduced into both OspA and LA-2 based on computational predictions on …

Inhibition Of Nuclear Factor-Kappa B Enhances The Tumor Growth Of Ovarian Cancer Cell Line Derived From A Low-Grade Papillary Serous Carcinoma In P53-Independent Pathway, Xue Xiao, Gong Yang, Peng Bai, Shunping Gui, Tri M. Bui Nguyen, +8 Additional Authors

Biochemistry and Molecular Medicine Faculty Publications

Background: NF-kB can function as an oncogene or tumor suppressor depending on cancer types. The role of NF-kB in low-grade serous ovarian cancer, however, has never been tested. We sought to elucidate the function of NF-kB in the low-grade serous ovarian cancer.

Methods: The ovarian cancer cell line, HOC-7, derived from a low-grade papillary serous carcinoma. Introduction of a dominant negative mutant, IkBαM, which resulted in decrease of NF-kB function in ovarian cancer cell lines. The transcription ability, tumorigenesis, cell proliferation and apoptosis were observed in derivative cell lines in comparison with parental cells.

Results: Western blot analysis indicated increased …

Blood-Tissue Barriers And Autoantibodies In Neurodegenerative Disease Pathogenesis: An Approach To Diagnostics And Disease Mechanism, Eric Luria Goldwaser

Graduate School of Biomedical Sciences Theses and Dissertations

Brain homeostasis can be affected in a number of ways that lead to gross anatomical, cellular, and molecular disturbances giving rise to diseases like Alzheimer’s disease (AD) and related dementias. Unfortunately, the mechanistic pathoetiology of AD’s hallmark features of cerebral amyloid plaque buildup and neuronal death are still disputed. Using human brain AD sections, immunohistochemistry experiments revealed internalized surface proteins, co-localized to an expanded lysosomal compartment. Other stains for amyloid-β1-42 (Aβ42) and various immunoglobulin (Ig) species displayed them leaking out of the cerebrovasculature through a dysfunctional blood-brain barrier (BBB), binding to neurons in the vicinity, and localizing to intracellular vesicles …

Investigating The Roles Of Δnp63 As A Suppressor Of Migration, Invasion, And Metastasis, Ramon E. Flores Gonzalez

Dissertations & Theses (Open Access)

Cancer is one of the leading causes of death and disease in the world. Considerable resources are spent to study and understand cancer, with the hope of developing new treatments and eventually cures that will help millions of people. Efforts to understand cancer are hindered by its inherent complexity and instability. Nonetheless, understanding the basics of tumor development and progression are the key to focused on studying the role of ΔNp63 in cancer, a p53 family member known to be involved in epithelial development, microRNA biogenesis, and stem cell maintenance. Using the strength of in vivo mouse models, we found …

Countercurrent Chromatography Fractions Of Plant Extracts With Anti-Tuberculosis Activity, Douglas Armstrong, Nathan C. Krause, Drew Frey, J. Brent Friesen, Baojie Wan, Jordan Gunn, Scott Franzblau

Faculty Scholarship – Chemistry

Samples of numerous plant species were received from the southwestern part of the USA, from Richard Spjut, and plant samples were collected here in Illinois. All were extracted with typical solvents, giving crude residues, some of which were subjected to chromatographic methods. Some of the crude residues and some of the fractions were tested for anti-tuberculosis activity and/or antibacterial activity.

In a general way, bioactive natural products are dealt with very well by Liang & Fang. More specifically, the southwestern part of the United States has a large variety of indigenous plants many of which have not been investigated for …

¬¬Define The Epigenetic Profiles And Subtype-Specific Genes Of Breast Cancer, Wenqian Li

Dissertations & Theses (Open Access)

Molecular profiling has identified 5 distinct subtypes of breast cancer, luminal A, luminal B, HER2-enriched, basal-like, and claudin-low breast cancer. These 5 subtypes correlate with hormone response, patient prognosis, and response to therapy. Although steady state gene expression patterns have been explored using expression microarrays, very little is known about the initial, disease-driving transcriptional changes in these cancers or epigenetic changes associated with the differential gene expression signatures. Defining these changes may provide new insights into the mechanisms by which these subtypes arise, as well as new avenues for breast cancer prevention, diagnosis, and treatment. Using Chromatin Immunoprecipitation sequencing and …

Circumventing Cisplatin Resistance In Ovarian Cancers Through Reactivation Of P53 By Non-Cross-Resistant Platinum Analogs, Michelle Martinez-Rivera

Dissertations & Theses (Open Access)

Abstract

CIRCUMVENTING CISPLATIN RESISTANCE IN OVARIAN CANCERS THROUGH REACTIVATION OF P53 BY NON-CROSS-RESISTANT PLATINUM ANALOGS

Michelle Martinez-Rivera, B.S.

Advisory Professor: Zahid H. Siddik, Ph.D.

Cisplatin (cis-Pt), an anticancer platinum (Pt) drug, is used widely in the treatment of several malignancies, such as ovarian cancer. This Pt compound induces DNA damage, which results in p53 activation through post-translational modifications, mainly phosphorylation, culminating in execution of programmed cell-death. However, despite initial therapeutic response to cis-Pt, clinical resistance to this drug emerges leading to disease progression. Pt-resistance phenotypes have been associated with dysfunction in the p53 signaling pathway. Therefore, an effort to understand …

A Caenorhabditis Elegans Model To Detect Developmental Teratogens With Adverse Reproductive And Metabolic Effects, Alexis Ann Killeen

USF Tampa Graduate Theses and Dissertations

Obesity is a current epidemic in the United States. Greater than one third of adults and nearly one fifth of children are classified as obese (Ogden, 2014). Increased awareness of this problem has triggered interest into the potentially causal agents in our environment. These agents, termed obesogens, are a type of teratogen hypothesized to work during embryonic development by programming offspring to store excess fat. A type of teratogen, obesogens are also typically reprotoxic, affecting germline development and embryonic viability. These noxious teratogens span a variety of different chemical classes including estrogen mimics, metals and metalloids, biocides, and even voluntary …

A Biochemical Approach To Define The Interactome For Calpain2 In Endothelial Cells, Dorothy E. Croall Phd, Jordan R. Hoffman Ms, Calvin Ph Vary Phd, Sharon L. Ashworth Phd, Billy R. Chase, Jay Beaudoin Ii

Molecular & Biomedical Sciences Faculty Scholarship

Current repositories for protein-protein interactions and high throughput screening methods focus on individual gene products and do not consider the significance of calcium induced conformational changes. These limitations suggest the need for alternative strategies to better define the calpain2 interactome. Affinity capture coupled with LC-MS/MS and proteomic analysis of the recovered proteins provides a powerful approach to identify protein-protein interactions for the heterodimeric calpain2. CAPN2 (rat) was modified to be catalytically incompetent (C105A) and fused with a C-terminal 15 residue peptide optimized for biotinylation by the biotin protein ligase, BirA. The resulting CAPN2*, heterodimerized with truncated CAPNS1, was purified from …

Cell Cycle Arrest By Tgfß1 Is Dependent On The Inhibition Of Cmg Helicase Assembly And Activation, Brook Samuel Nepon-Sixt

USF Tampa Graduate Theses and Dissertations

Tumorigenesis is a multifaceted set of events consisting of the deregulation of several cell-autonomous and tissue microenvironmental processes that ultimately leads to the acquisition of malignant disease. Transforming growth factor beta (TGFß) and its family members are regulatory cytokines that function to ensure proper organismal development and the maintenance of homeostasis by controlling cellular differentiation, proliferation, adhesion, and survival, as well as by modulating components of the cellular microenvironment and immune system. The pleiotropic control by TGFß of these cell intrinsic and extrinsic factors is intimately linked to the prevention of tumor formation, the specifics of which are dependent on …

Regulation Of Phosphatase And Tensin Homolog Expression And Activity By Transforming Growth-Factor Beta In The Trabecular Meshwork Cells: Implications For Primary Open Angle Glaucoma, Nikoleta Tellios

Electronic Thesis and Dissertation Repository

Glaucoma is a multifactorial condition caused, in part, by fibrosis of the sieve-like trabecular meshwork (TM) tissue, which impedes drainage of aqueous humor (AH), leading to increased intraocular pressure and associated optic nerve damage and blindness. Fibrosis of the TM is mainly caused by the increased levels of active transforming growth factor-β 2 (TGFβ2) in the AH of glaucoma patients.

Previous reports have shown that TGFβ decreases the expression of Phosphatase and Tensin Homolog (PTEN) gene and that PTEN is a major regulator of ECM deposition. In this study we investigate the regulation of PTEN protein expression and …