Biochemistry, Biophysics, and Structural Biology Commons^™

Myxobacteria Versus Sponge-Derived Alkaloids: The Bengamide Family Identified As Potent Immune Modulating Agents By Scrutiny Of Lc-Ms/Elsd Libraries., Tyler A. Johnson, Johann Sohn, Yvette M. Vaske, Kimberly N. White, Tanya L. Cohen, Helene C. Vervoort, Karen Tenney, Frederick A. Valeriote, Leonard F. Bjeldanes, Phillip Crews

Natural Sciences and Mathematics | Faculty Scholarship

A nuclear factor-κB (NF-κB) luciferase assay has been employed to identify the bengamides, previously known for their anti-tumor activity, as a new class of immune modulators. A unique element of this study was that the bengamide analogs were isolated from two disparate sources, Myxococcus virescens (bacterium) and Jaspis coriacea (sponge). Comparative LC-MS/ELSD and NMR analysis facilitated the isolation of M. viriscens derived samples of bengamide E (8) and two congeners, bengamide E' (13) and F' (14) each isolated as an insperable mixture of diastereomers. Additional compounds drawn from the UC, Santa Cruz repository allowed expansion of the structure activity relationship …

Development Of A Chaperonin Activity Assay, Timothy P. Brown, Danielle Y. Martinez, Ricardo A. Bernal

COURI Symposium Abstracts, Summer 2012

Chaperonin-mediated protein folding by the Escherichia coli GroEL/GroES complex is pivotal in preventing premature aggregation and improper folding of proteins. In fact E. coli that have had the groE operon knocked out are not able to survive. The explicit details of facilitated folding in the interior of the chaperonin complex remain relatively unknown, although it is understood that ATP hydrolysis is essential to the chaperonin protein-folding cycle. The goal of this project is to develop an assay to detect the protein folding activity of chaperonins. ATP hydrolysis reactions allow chaperonins to undergo a series of conformational changes that alter the …

Protein Expression Analysis Of Cell Lines Derived From Drug Resistant Tumors, Paloma Valenzuela, Karla Parra, Natzidielly Lerma, Courtney Becerril, Eduardo Ramirez, Irving Miramontes, Howard West, Cynthia Rodriguez, Yang Li, Jianying Zhang, Giulio Francia

COURI Symposium Abstracts, Summer 2012

There are a number of effective treatments for breast cancer, including chemotherapy and targeted strategies such as the use of the Her-2 targeting drugs lapatinib and trastuzumab. However, in a number of cases, tumors that initially respond to therapy eventually develop drug resistance, leading to the relapse of the disease. By studying protein levels in different drug resistant variants, we have observed two mechanisms by which drug resistance may develop. Thus, some Her-2 human breast cancer cells (e.g., MDA-MB-231H2N) treated with the clinically used trastuzumab agent, can escape therapy by shedding, or losing, the target Her-2 protein. Similarly, EMT-6 mouse …

Purification And Characterization Of Mycobacterium Tuberculosis Esx-1 Secreted Protein Esat-6, Sebastian Montalvo, Gouzhong Jiang, Jianjun Sun

COURI Symposium Abstracts, Summer 2012

Mycobacterium tuberculosis ESX-1 secreted protein ESAT-6 has been shown to contribute to the pathogenesis and virulence of M. tuberculosis. Despite many studies, little is known about how this protein interacts with the host’s membrane and evades the host’s defense mechanism. In this project, we successfully purified and characterized of ESAT-6 wild type (E6-WT) and ESAT-6 mutant with either a single (E6-S) or double (E6-D) amino acid mutation that are supposed to diminish the membrane-interacting activity of ESAT-6. The methods used in our project include: cloning in plasmid vectors, cloning in PCR products with T-Vectors, chemical transformation of E. coli, affinity …

Phytochemical Intervention In Protein Misfolding Linked To Neurodegenerative Diseases, Acantha A. Abdulla, Emmanuel Zubia, Mahesh Narayan

COURI Symposium Abstracts, Summer 2012

Protein Disulfide Isomerase (PDI) is the chief protein folding chaperone found in the rough endoplasmic reticulum. While folding maturation is accelerated in the presence of PDI, S-nitrosylated PDI, arising from reactive oxygen species stress, causes accumulation of misfolded debris. S-nitrosylated PDI has been found in the neuronal cells of victims of both Alzheimer’s and Parkinson’s diseases. Our objective is to introduce antioxidant-rich extracts from the Azadirachta Indica (Neem) leaf to protect PDI from reactive oxygen species (S-nitrosylation). We anticipate that Neem extracts can be neuroprotective and serve as a prophylactic for mitigating the incidence of reactive-oxygen species related neurodegenerative diseases. …

Cytotoxic Effects Of Creosote (Larrea Tridentata) Plant Extracts On Human Lymphoma Cells Lines, Emanuel Cordero, Yahaira Santiago, Carolina Lema, Armando Varela-Ramirez, Renato J Aguilera

COURI Symposium Abstracts, Summer 2012

Larrea tridentata also known as Creosote is a North American shrub, whose leave extracts are proposed to contain anti-cancer properties. The main metabolite in this plant is the nordihydroguaiaretic acid (NDGA) has been shown to have anti-neoplastic, anti-viral and anti-inflammatory properties. NDGA is a strong anti-oxidant that can scavenge or inhibit reactive oxygen spices production, stimulate nitric oxide production, increase immune function, enhance central nervous system function, and prevent cardiovascular or other diseases. Although extracts from this plant are currently used in Mexico as an herbal medicine, the Food and Drug Administration and Health Canada have issued health hazard warming …

In Vitro Evaluation Of Human Her2-Positive Breast Cancer Cells, Karla Parra, Howard West, Paloma Valenzuela, Eduardo D. Ramirez, Natzidielly Lerma, Courtney Becerril, Irving Miramontes, Cynthia M. Rodriguez, Guido Bocci, Giulio Francia

COURI Symposium Abstracts, Summer 2012

One in four cases of breast cancer shows over expression of the Her-2 protein, and there is an urgent need to improve therapies for this disease. We have evaluated two Her-2 positive human breast cancer cell lines, MDA-231H2N and HCC1419. MDA-231 human breast cancer cell, which are Her-2 negative, were used as a control. The cell lines were evaluated in vitro for their relative growth rate, their growth under reduced (i.e. 3% serum) conditions, or as spheroid cultures, and for their sensitivity to drugs such as ceramide analogs and tyrosine kinase inhibitors (i.e. CLM3, CLM29, and CLM94) that are being …

Identification Of Two Functional Sumo Interacting Motifs (Sims) In The Non-Structural Protein Of Influenza A Virus Ns1a, Lorellie M. Lewis, German Rosas-Acosta

COURI Symposium Abstracts, Summer 2012

We have previously shown that the non-structural protein of Influenza A virus NS1A is post-translationally modified by conjugation of the Small Ubiquitin-like MOdifier (SUMO). Our functional studies have also demonstrated that SUMO conjugation (SUMOylation) affects NS1A's ability to neutralize the cellular interferon response of the host cell and other functions associated to NS1A. Importantly, we have also shown that influenza infections trigger a global increase in cellular SUMOylation and have recently demonstrated that such an increase is mediated by NS1A. This observation led us to investigate the molecular mechanism endowing NS1A with the ability to increase global cellular SUMOylation. Sequence …

Generation And Characterization Of Sequence-Independent Sumoylated Lens Epithelium-Derived Growth Factor (Ledgf) Protein P75 Chimeras., Alfredo Roman, Denisse Gutierrez, Manuel Llano

COURI Symposium Abstracts, Summer 2012

LEDGF/p75 is a transcriptional coactivator required for efficient lentiviral cDNA integration. LEDGF/p75 has been found to be SUMOylated at four lysine residues, and mutation of these residues to arginine inhibits LEDGF/p75 SUMOylation and impairs the HIV-cofactor activity of this protein. However, we still do not understand how SUMOylation influences the HIV cofactor activity of LEDGF/p75. It has been demonstrated that attachment of SUMO to transcription factors in a non-sequence specific fashion recapitulates the effect of sequence-specific SUMOylation on the activity of these proteins. These results have suggested that is SUMO per se and not the modification of the target protein …

The Synthesis And Structural Characterization Of Metal Oxide Nanoparticles Having Catalytic Applications, Stacey Janel Smith

Theses and Dissertations

Nanotechnology is blossoming into one of the premiere technologies of this century, but the key to its progress lies in developing more efficient nanosynthesis methods. Variations in synthetic technique, however, can cause variations in size, structure, and surface characteristics, thereby altering the physical properties and functionality of the particles. Careful structural characterizations are thus essential for understanding the properties and appropriate applications for particles produced by new synthetic techniques.In this work, a new ‘solvent-deficient’ method is presented for the synthesis of an unprecedentedly wide range of metal oxide nanomaterials including at least one metal oxide from each group in Groups …

Inhibitory Effects Of Novel Immucillin Analogues On Borrelia Burgdorferi Bgp Nucleosidase, Christian Guerrero, Seth Eidemiller, Ken Cornell

IDeA Network for Biomedical Research Excellence (INBRE)

The pathogenic spirochaete Borrelia burgdorferi causes Lyme disease and is transmitted by deer ticks when they feed. Lyme disease is multisystemic—it adversely affects the heart, joints, and skin. Recent studies demonstrate that B. burgdorferi possesses three methylthioadenosine/Sadenosylhomocysteine (MTA/SAH) nucleosidases essential for the catabolic breakdown of both MTA and SAH. Both MTA and SAH are by-products of major pathways involving Sadenosylmethionine (SAM) and are kept at low micromolar concentrations due to their inhibitory activity.

This project examined the effect of transition state inhibitors on the surface binding Borrelia glycosaminoglycanbinding protein (Bgp) nucleosidase using recombinant Bgp and whole-cell B. burgdorferi activity assays. …

Identification Of Persistent Long Range Interactions In GA95 And GB95 Through Thermal Unfolding Simulations, Milen Redai Tesfamariam

Chemistry & Biochemistry Theses & Dissertations

For over five decades, different experiments have been performed to research how proteins attain their native three dimensional structures. However, the folding problem continues to be a puzzle in modern science. The design of two proteins that have maximal sequence identity but different folds and functions is one method that is being used to study the relationship between protein structure and amino acid sequence. In particular, mutant proteins of Streptococcus protein G, G_A and G_B, have 95% sequence identity and a 3a helix fold and β4/a fold, respectively. Molecular dynamics simulations of G_A95 …

Centromeric Heterochromatin Assembly In Fission Yeast-Balancing Transcription, Rna Interference And Chromatin Modification, Benjamin Alper, Brandon R. Lowe, Janet F. Partridge

Chemistry & Physics Faculty Publications

Distinct regions of the eukaryotic genome are packaged into different types of chromatin, with euchromatin representing gene rich, transcriptionally active regions and heterochromatin more condensed and gene poor. The assembly and maintenance of heterochromatin is important for many aspects of genome control, including silencing of gene transcription, suppression of recombination, and to ensure proper chromosome segregation. The precise mechanisms underlying heterochromatin establishment and maintenance are still unclear, but much progress has been made towards understanding this process during the last few years, particularly from studies performed in fission yeast. In this review, we hope to provide a conceptual model of …

Dietary Analysis And Epigenetic Comparisons Of Drosphila Melanogaster Through Multiple Generations, Xu (Kevin) F. Zheng

Holster Scholar Projects

Nutrition and the lack thereof has been a demanding issue in both the undeveloped and developed country. The unbalance diet between healthy and unhealthy diet has led to many of the problems presented in society such as obesity, heart disorders, and loss of longevity. It is now known that calorie may not provide the most accurate information in terms of the lifestyle and the well-being of humans. Rather the composition of the calorie is what becomes integral in people's understanding of the effect of food physiologically. With this knowledge, we seek to use a Drosophila melanogaster as a model organism …

Caehnorhabditis Elegans: A Low-Cost In Vivo Animal Model For Efficacy Studies Of Novel Antibiotics, Rylee Ann Gregory

Undergraduate Honors Capstone Projects

Since the 1940s, antibiotics have greatly reduced the adverse effects of infectious diseases caused by microbes. However, due to excessive, and often incorrect, use of known antibiotics, many organisms have adapted antibiotic resistance. Currently, over 70% of known infectious bacteria are resistant to at least one antibiotic. In the U.S. , 90,000 deaths occur each year due to infection by bacteria resistant to antibiotics. This number has increased by nearly 75,000 in the last 20 years. It is necessary, therefore, to continue developing new antibiotics in an effort to keep up with increasing antibiotic resistance. Traditional in vitro and whole …

Screening Synthetic Combinatorial Protein Libraries For Changes In Calmodulin Binding Specificity, Donald E. Dick

Kaleidoscope

No abstract provided.

Starch Phosphates: A Means To Manipulate Biofuel Production, Travis M. Bridges

Kaleidoscope

No abstract provided.

The Role Of Argininosuccinate Synthase Serine 328 Phosphorylation In Nitric Oxide Production, Ricci Haines

USF Tampa Graduate Theses and Dissertations

Until recently, the main mechanism of argininosuccinate synthase (AS) regulation was described to exist mainly at the level of transcription. Transcriptional regulation of AS has been shown to be coordinate with eNOS in response to shear stress, hypoxia, tumor necrosis factor á (TNF-á), and PPAR ã agonist troglitizone. However, it is now understood that one level of NO regulation is cellular control of arginine availability to eNOS via post-translational modifications of AS such as phosphorylation. The purpose of this investigation was to determine under what conditions AS is phosphorylated at S328, identify the pathway that AS phosphorylation at S328 plays …

Live-Cell Monitoring Of Periodic Gene Expression In Synchronous Human Cells Identifies Forkhead Genes Involved In Cell Cycle Control, Gavin D. Grant, Joshua Gamsby, Viktor Martyanov, Lionel Brooks, Lacy K. George, J. Matthew Mahoney, Jennifer J. Loros, Jay C. Dunlap, Michael L. Whitfield

Dartmouth Scholarship

We developed a system to monitor periodic luciferase activity from cell cycle-regulated promoters in synchronous cells. Reporters were driven by a minimal human E2F1 promoter with peak expression in G1/S or a basal promoter with six Forkhead DNA-binding sites with peak expression at G2/M. After cell cycle synchronization, luciferase activity was measured in live cells at 10-min intervals across three to four synchronous cell cycles, allowing unprecedented resolution of cell cycle-regulated gene expression. We used this assay to screen Forkhead transcription factors for control of periodic gene expression. We confirmed a role for FOXM1 and identified two novel cell cycle …

Mda-7/Il-24; A Promising Cancer Therapeutic Agent, Hossein Hamed

Theses and Dissertations

Glioblastoma multiforme (GBM) is an aggressive cancer that affects millions of patients per year. Conventional therapies combining chemotherapeutic agents with radiation can only extend survival by a few months; therefore, there is a dire need for an effective means of treating this deadly disease. Melanoma differentiation-associated gene-7/interleukin-24 (mda-7/IL-24), currently in the early stages of FDA pre-IND drug trials, has proven to be an effective cancer specific cytokine, able to trigger the onset of mitochondrial dysfunction and/or autophagy. GBM’s have mutations that often result in the activation of cytoprotective cell signaling pathways, preventing cancer therapeutics and even MDA-7/IL-24 treatments from being …

Sigma Factor N (Σn): A Novel Regulator Of Extreme Acid Resistance In Enterohemorrhagic E. Coli O157:H7, Pamela Ann Fay

USF Tampa Graduate Theses and Dissertations

Extreme acid resistance contributes to the successful transmission of enterohemorrhagic E. coli (EHEC) through acidic food matrices and the stomach, allowing it to gain access to the intestine and elicit disease in humans. Alternative sigma factor N (σ^N, encoded by rpoN) was previously identified as a novel regulator of extreme acid resistance in EHEC. This study investigated the role for σ^N and co-expressed products of the rpoN operon in the acid resistance phenotype of EHEC. The results revealed that σ^N primarily controls acid resistance through repression of the glutamate-dependent acid resistance (GDAR) system through control …

(1)H, (15)N And (13)C Backbone Resonance Assignments Of The Kelch Domain Of Mouse Keap1., Elio Cino, Jingsong Fan, Daiwen Yang, Wing-Yiu Choy

Biochemistry Publications

Kelch-like ECH-associated Protein 1 (Keap1) is a multi-domain protein that functions as an inhibitor of the transcription factor nuclear factor E2-related factor 2 (Nrf2) in the cellular response to oxidative stress. Under normal conditions, Keap1 binds to Nrf2 via its C-terminal Kelch domain and the interaction ultimately leads to the ubiquitin-dependent degradation of Nrf2. It has been proposed that designing molecules to selectively disrupt the Keap1-Nrf2 interaction can be a potential therapeutic approach for enhancing the expression of cytoprotective genes. Here, we reported the (1)H, (13)C, and (15)N backbone chemical shift assignments of the Kelch domain of mouse Keap1. Further, …

Systematics Of Harrisia (Cactaceae), Alan R. Franck

USF Tampa Graduate Theses and Dissertations

The genus Harrisia Britton (Cactaceae) comprises species of columnar cacti that are united by a unique seed morphology. The species range in form from prostrate shrubs to large trees and are native to South America and the Caribbean region. Harrisia is placed in an unresolved position within subtribe Trichocereinae of tribe Cereeae of subfamily Cactoideae. Relationships among the species within Harrisia are also poorly understood. In this study, several species of Harrisia were sequenced for as many as seven different regions of nuclear and plastid DNA. Species in the Caribbean were also examined with amplified fragment length polymorphisms. The morphology …

Association Of A Multi-Synthetase Complex With Translating Ribosomes In The Archaeon Thermococcus Kodakarensis, Medha Raina, Sara Elgamal, Thomas J. Santangelo, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

In archaea and eukaryotes aminoacyl-tRNA synthetases (aaRSs) associate in multi-synthetase complexes (MSCs), however the role of such MSCs in translation is unknown. MSC function was investigated in vivo in the archaeon Thermococcus kodakarensis, wherein six aaRSs were affinity co-purified together with several other factors involved in protein synthesis, suggesting that MSCs may interact directly with translating ribosomes. In support of this hypothesis, the aminoacyltRNA synthetase (aaRS) activities of the MSC were enriched in isolated T. kodakarensis polysome fractions. These data indicate that components of the archaeal protein synthesis machinery associate into macromolecular assemblies in vivo and provide the potential …

Investigation Into The Mechanism Of Salicylate-Associated Genotypic Antibiotic Resistance In Staphylococcus Aureus, Nada Salah Helal

USF Tampa Graduate Theses and Dissertations

Growth of Staphylococcus aureus with the NSAID salicylate increases phenotypic resistance (SAPAR), and the frequency at which heritable resistance occurs to various antibiotics (SAGAR). This study describes the effect of salicylate on heritable and phenotypic resistance to a set of antibiotics for laboratory and multi-drug resistant strains of S. aureus and investigates the link between resistance and SAGAR. Drug gradient plates were used to determine phenotypic resistance to antibiotics targeting DNA replication, transcription, translation and the cell wall in the presence or absence of salicylate. To measure heritable resistance, mutation frequencies were determined for each antibiotic in the presence and …

Aspirin But Not Ibuprofen Use Is Associated With Reduced Risk Of Prostate Cancer: A Plco Study., F. M. Shebl, L. C. Sakoda, A. Black, J. Koshiol, G. L. Andriole, R. Grubb, T. R. Church, D. Chia, C. Zhou, L. W. Chu, W. Y. Huang, U. Peters, V. A. Kirsh, N. Chatterjee, M. F. Leitzmann, R. B. Hayes, A. W. Hsing

Biochemistry and Molecular Medicine Faculty Publications

Background:

Although most epidemiological studies suggest that non-steroidal anti-inflammatory drug use is inversely associated with prostate cancer risk, the magnitude and specificity of this association remain unclear.

Methods:

We examined self-reported aspirin and ibuprofen use in relation to prostate cancer risk among 29 450 men ages 55–74 who were initially screened for prostate cancer from 1993 to 2001 in the Prostate, Lung, Colorectal, and Ovarian Cancer Screening Trial. Men were followed from their first screening exam until 31 December 2009, during which 3575 cases of prostate cancer were identified.

Results:

After adjusting for potential confounders, the hazard ratios (HRs) of …

Role Of The Nla6s And Nla28s Histidine Kinases In Fruiting Body Development Of Myxococcus Xanthus, Zaara Sarwar

Biology - Dissertations

The complex life cycle of Myxococcus xanthus makes it a model organism for studying multicellular developmental processes in bacteria. In response to adverse environmental conditions, M. xanthus aggregates and forms multicellular structures known as fruiting bodies. Two component signal transduction systems (TCS) are widely used by bacteria to detect and respond to environmental cues by regulating large-scale changes in gene expression. They contain a histidine kinase sensor that detects environmental cues and a response regulator that modulates cellular processes. Two key regulators of the early stages of fruiting body development are the Nla6S/Nla6 and Nla28S/Nla28 TCSs. The response regulators of …

Minor Fibrillar Collagens, Variable Regions Alternative Splicing, Intrinsic Disorder, And Tyrosine Sulfation, Ming Fang, Reed B. Jacob, Owen Mcdougal, Julia Oxford

Biomedical Research Institute Publications and Presentations

Minor fibrillar collagen types V and XI, are those less abundant than the fibrillar collagens types I, II and III. The alpha chains share a high degree of similarity with respect to protein sequence in all domains except the variable region. Genomic variation and, in some cases, extensive alternative splicing contribute to the unique sequence characteristics of the variable region. While unique expression patterns in tissues exist, the functions and biological relevance of the variable regions have not been elucidated. In this review, we summarize the existing knowledge about expression patterns and biological functions of the collagen types V and …

A Proteomic Analysis Of Differentiation In The Mammary Epithelium, Laura Therese Strand

Master's Theses

While a great deal is known about the changing hormonal environment and the structural development of the mammary gland from pregnancy to lactation, very little is known about the molecular mechanisms governing differentiation of the mammary epithelium into a milk-secreting phenotype. It is important to acknowledge the diversity among the mammary glands of different species in order to better understand applications in human health and the dairy industry. In this study, we examined global protein expression during two states of differentiation in mammary epithelial cells from two species: in vitro proliferating and differentiated MAC-T cells (a bovine immortal cell-line), and …

Development Of A Protocol To Measure Gene Expression In The Mouse Tibia, Daniel Hoover

Biomedical Engineering

Numerous molecular factors active in bone tissue direct fracture repair and remodeling which can be altered by disease conditions such as Peripheral Arterial Disease (PAD) and Osteoporosis. Methods of molecular biology are commonly applied to investigate the expression and role of these molecular factors. This project presents a robust three-step protocol for examining gene expression in the mouse tibia. The protocol begins with isolating RNA from a flash frozen tibia sample. The isolated RNA is reverse transcribed into cDNA. Finally, PCR is performed to indentify expressed genes. Establishing this protocol will allow further research into the mechanisms of bone remodeling …