Molecular Biology Commons^™

Structure/Function Analysis Of Fega And Fhua In Bradyrhizobium Sp., Alexander James Herd

Senior Honors Projects, 2010-2019

Bradyrhizobium japonicum is a Gram-negative soil bacterium commonly known for its agriculturally significant mutualistic relationship with soybean. In this symbiosis, the bacteria and plant undergo complex molecular signaling characterized by sent and received signals resulting in the formation of infection threads and root nodules. This research aimed to compare two related bacterial outer membrane proteins, FegA and FhuA, associated with the molecular signaling between the bacteria and plant. Previous work has led to the hypothesis that the N-terminal domain (NTD) of FegA in B. japonicum is needed for a functional symbiosis to occur. Recombinant bacterial strains expressing altered FegA proteins …

Toward An Enzyme-Coupled, Bioorthogonal Platform For Methyltransferases: Probing The Specificity Of Methionine Adenosyltransferases, Tyler D. Huber

Theses and Dissertations--Pharmacy

Methyl group transfer from S-adenosyl-l-methionine (AdoMet) to various substrates including DNA, proteins, and natural products (NPs), is accomplished by methyltransferases (MTs). Analogs of AdoMet, bearing an alternative S-alkyl group can be exploited, in the context of an array of wild-type MT-catalyzed reactions, to differentially alkylate DNA, proteins, and NPs. This technology provides a means to elucidate MT targets by the MT-mediated installation of chemoselective handles from AdoMet analogs to biologically relevant molecules and affords researchers a fresh route to diversify NP scaffolds by permitting the differential alkylation of chemical sites vulnerable to NP MTs that are unreactive to …

Cyclophilin A As A Molecular Switch Regulating Prolactin Receptor Mediated Signaling, Mammary Tumorigenesis And Metastasis, Shawn Hakim

Theses and Dissertations

Prolactin (PRL) and its receptor (PRLr) have been implicated in the development and progression of human breast cancer. PRL activates its receptor and induces activation of proximal Janus kinase 2 (Jak2) for signal transduction. Here, we sought to determine the role of PRLr-associated peptidyl-prolyl isomerase, cyclophilin A (CypA), in modulating structure/function relationships of the PRLr. It was demonstrated that CypA mediated PRL-induced conformational change of the CFP- and -YFP tagged forms of the PRLr cytoplasmic-tail, whereas CypA inhibition by NIM811 (N-methyl-4-isoleucine cyclosporin) or knockdown blocked the conformational change of the PRLr assessed by Fluorescence Resonance Energy Transfer (FRET) signal or …

Preconditioning Of The Tumor Microenvironment By Means Of Low Dose Chemotherapies For An Effective Immunotherapy Of Breast Cancer, Hussein F. Aqbi

Theses and Dissertations

Breast cancer mortality is mainly due to distant recurrence of the disease arising from dormant tumor cells established by cancer therapies. Patients who initially respond to cancer therapies often succumb to distant recurrence of the disease. It is not clear why people with the same type of breast cancer respond to treatments differently; some escape from dormancy and relapse earlier than others. In addition, some tumor clones respond to immunotherapy while others do not. We investigated how autophagy plays a role in accelerating or delaying recurrence of neu overexpressing mouse mammary carcinoma (MMC) following adriamycin (ADR) treatment, and in affecting …

In Vivo Assessment Of Safranal’S Novel Therapeutic Effects On Chemically Induced Hepatic Neoplasia, Ameera Ali M. Almansoori

Biology Theses

Chronic liver insult leads to fibrosis, which often ends up causing cirrhosis and most of the time that progresses into hepatic neoplasms (early Hepatocellular carcinoma; HCC). HCC is the fifth most common cancer, and the third cause of cancerrelated deaths. Chemotherapy is the most common treatment of cancer patients. HCC is however, chemo-resistant, and the side-effects of chemotherapy are typically exhausting to the patient. Sorafenib is the only anti-HCC drug approved by the U.S Food and Drug Administration. It is a multikinase inhibitor that blocks tumor cells proliferation and angiogenesis. Although sorafenib is successful treating early and mid HCC lesions, …

Transcriptional Profiling Identifies Potential Genes Associated With Multiple Biotic Stresses In Arabidopsis, Hibatullah Mustafa Al Ashram

Biology Theses

Plants are exposed to many environmental stresses that affect their growth and development. These stresses include biotic stresses (organisms) and abiotic stresses (drought and salinity). Plants respond to these stresses by transcriptional reprogramming and different signaling pathways. Arabidopsis thaliana had shown great sensitivity to the biotic stress: Botrytis cinerea, Alternaria brassicicola, Pseudomonas syringae and the herbivore insect Pieris rapae. The overall goal is to identify common regulated genes that respond to all these biotic stresses to ultimately improve plant stress tolerance in Arabidopsis. The specific aims are to: (1) determine the regulated genes in response to an …

Adapting Cell-Free Protein Synthesis As A Platform Technology For Education, Grace W. Chu, Max Z. Levine, Nicole E. Gregorio, Javin P. Oza

STAR Program Research Presentations

Cell-free protein synthesis (CFPS) has emerged as an enabling biotechnology for research and biomanufacturing as it allows for the production of protein without the need for a living cell. Applications of CFPS include the construction of libraries for functional genomics and structural biology, the production of personalized medicine, and the expression of virus-like particles. The absence of a cell wall provides an open platform for direct manipulation of the reaction conditions and biological machinery. This project focuses on adapting the CFPS biotechnology to the classroom, making a hands-on bioengineering approach to learning protein synthesis accessible to students grades K-16 through …

Direct Quantification Of Deubiquitinating Enzyme Activity In Single Intact Cells, Nora Safabakhsh

LSU Doctoral Dissertations

Challenges in drug efficacy occur during the treatment of most types of cancer due to the heterogeneity of the tumor microenvironment. This has led to the development of personalized medicine. Due to the clinical success of the proteasome inhibitors Bortezomib and Carfilzomib in treatment of multiple myeloma, interest has shifted towards molecularly-targeted chemotherapeutics for ubiquitin-proteasome system (UPS). Deubiquitinating enzymes (DUBs) are an essential part of this pathway which have been found to promote Bortezomib resistance in multiple myeloma patients. Unfortunately, there is a lack of specific, high throughput biochemical assays to characterize DUB activity in patient samples before and after …

Engineering Bioluminescent Sensors Of Cyclic Amp To Study Opioid Signaling, Alexander L. Tesmer, Alexander R. French, Mathew Tantama

The Summer Undergraduate Research Fellowship (SURF) Symposium

Opioids are small signaling molecules which bind to opioid receptors on the surface of cells. The kappa opioid receptor (KOR) is one of three major types of opioid receptors found in human neurons. When an opioid binds to a KOR, a variety of biochemical signaling pathways are activated inside the cell. Each of these pathways are associated with different physiological effects of KOR activation. The production of a small signaling molecule, cyclic adenosine monophosphate (cAMP), is known to be inhibited during KOR activation of the analgesic (pain-killing) signaling pathway. The ability to interrogate the individual responses of KOR signaling pathways …

Transcriptomics Of Learning, Pablo Iturralde

Theses

Learning is a basic and important component of behavior yet we have very little empirical information about the interaction between mechanisms of learning and evolution. In our work, we are testing hypotheses about the neurogenetic mechanisms through which animal learning abilities evolve. We are able to test this directly by using experimentally evolved populations of flies, which differ in learning ability. These populations were previously evolved within the lab by creating worlds with different patterns of change following theoretically predicted effects on which enhanced learning will evolve. How has evolution acted to modulate genes and gene expression in the brain …

Mapping Molecular Datasets Back To The Brain Regions They Are Extracted From: Remembering The Native Countries Of Hypothalamic Expatriates And Refugees, Arshad Khan, Alice Grant, Anais Martinez, Gully Burns, Brendan Thatcher, Vishwanath Anekonda, Benjamin Thompson, Zachary Roberts, Daniel Moralejo, James Blevins

Selected Works Temporary Series

This article, which includes novel unpublished data along with commentary and analysis,focuses on approaches to link transcriptomic, proteomic, and peptidomic datasets mined frombrain tissue to the original locations within the brain that they are derived from using digital atlasmapping techniques. We use, as an example, the transcriptomic, proteomic and peptidomicanalyses conducted in the mammalian hypothalamus. Following a brief historical overview, wehighlight studies that have mined biochemical and molecular information from the hypothalamusand then lay out a strategy for how these data can be linked spatially to the mapped locations in acanonical brain atlas where the data come from, thereby allowing …

Mapping Molecular Datasets Back To The Brain Regions They Are Extracted From: Remembering The Native Countries Of Hypothalamic Expatriates And Refugees, Khan, Alice Grant, Anais Martinez, Gully Burns, Brendan Thatcher, Vishwanath Anekonda, Benjamin Thompson, Zachary Roberts, Daniel Moralejo, James Blevins

Departmental Papers (Biology)

This article, which includes novel unpublished data along with commentary and analysis,focuses on approaches to link transcriptomic, proteomic, and peptidomic datasets mined frombrain tissue to the original locations within the brain that they are derived from using digital atlasmapping techniques. We use, as an example, the transcriptomic, proteomic and peptidomicanalyses conducted in the mammalian hypothalamus. Following a brief historical overview, wehighlight studies that have mined biochemical and molecular information from the hypothalamusand then lay out a strategy for how these data can be linked spatially to the mapped locations in acanonical brain atlas where the data come from, thereby allowing …

Ketone Bodies And Signaling In Pancreatic Cancer Cell Lines, Kyla B. Buettner, Pankaj K. Singh, Surendra K. Shukla

Theses/Capstones/Creative Projects

Pancreatic cancer is the fourth leading cause of cancer-related deaths in the United States, and 95% of these cases are caused by PDAC (pancreatic ductal adenocarcinoma). Ketone bodies have previously been shown to decrease cell proliferation and cancer-induced cachexia. The molecular mechanism of ketone body-mediated growth inhibition of pancreatic cancer cells is not well understood. Research conducted thus far has not explored which molecular pathways are affected by ketone body treatment in pancreatic cancer cells. In the current study, the effect of the ketone body sodium hydroxybutyrate on the JAK-STAT and mTOR pathways and cell migration was explored. A decrease …

Optimizing Genetic Manipulation Of Methanogens Through Faster Cloning Techniques, Merrisa Jennings

Biological and Agricultural Engineering Undergraduate Honors Theses

Methanogenesis is the biological production of methane. Only anaerobic archaea known as methanogens are capable of such a metabolic feat. They have strict living conditions and substrate sources which determine their rate of metabolism. This is of particular importance from a greenhouse gas reduction perspective or biogas capturing perspective. One of the best ways to optimize methanogen methane production is via genetic manipulation. The current procedures are timely though, therefore a faster cloning processes should be developed. The objective of this study was to optimize a premade genetic transformation kit known as the Gibson Kit. The Gibson Kit was supposed …

Molecular Cloning And Functional Analysis Of Salt Overly Sensitive (Sos) Pathway Genes From Local Plants Of The United Arab Emirates, Mariam Obaid Saeed Abdalla Alzaabi

Biology Theses

Salinity is one of the major abiotic stresses that lead to extreme reduction in crop productivity worldwide. Increased soil and water salinity is a major issue in UAE’s agriculture. Some plants however have adapted to grow in high saline conditions, i.e. halophytes. Unfortunately, most crops are glycophytes and cannot tolerate high salinity. During salinity stress, expression of numerous genes is altered to protect the plants. Understanding the differences in gene structure between halophytes and glycophytes could provide insights into developing transgenic crops that can thrive in salinity conditions. The Salt Overly Sensitive (SOS) pathway is a key mechanistic cascade controlling …

Transcriptomic Analysis On Hepatocellular Carcinoma Model In Response To Safranal Treatment, Badriya Mahmood Ahmed Baig

Biology Theses

Unsolved riddle of cancer remains a major global health concern particularly in hepatocellular carcinoma (HCC) which is among the leading causes of cancer related death worldwide. Saffron (Crocus sativus L.) and its bioactive phytochemicals possess promising anti-cancer activities. One of these phytochemicals, safranal, has demonstrated potent anti-cancer activities against different types of cancers. Yet, its association with liver cancer remains under documented. This study investigated HCC response to safranal’s treatment via transcriptomic approach. Safranal was found to be involved in mitochondrial dysfunction, induced oxidative stress and increased reactive oxygen species (ROS) production leading to cell death. Interestingly, safranal has …

Anti-Colon Cancer Effect Of Origanum Majorana Essential Oil, Asma Nasser Rashed Alrashedi

Biology Theses

Plants have been shown to be an excellent source of new drugs, including anticancer agents. Origanum majorana commonly known as marjoram is a plant that is known to possess different therapeutic values including antioxidant andantimicrobial activities.

Our research team has previously tested the ethanolic extract of O. majorana on triple negative breast cancer and published the findings. The ethanolic extract promoted mitotic arrest at G2/M phase, induced apoptosis as well as inhibition of migration and metastasis. The promising potential of the ethanolic extract encouraged us to test the effects of O. majorana essential oil on human colon cancer cell lines. …

Quantum Confined Peptide Assemblies With Tunable Visible To Near-Infrared Spectral Range, Kai Tao, Zhen Fan, Leming Sun, Pandeeswar Makam, Zhen Tian, Mark Ruegsegger, Shira Shaham-Niv, Derek Hansford, Ruth Aizen, Zui Pan, Scott Galster, Jianjie Ma, Fan Yuan, Mingsu Si, Songnan Qu, Mingjun Zhang, Ehud Gazit, Junbai Li

Faculty & Staff Scholarship

Quantum confined materials have been extensively studied for photoluminescent applica- tions. Due to intrinsic limitations of low biocompatibility and challenging modulation, the utilization of conventional inorganic quantum confined photoluminescent materials in bio- imaging and bio-machine interface faces critical restrictions. Here, we present aromatic cyclo-dipeptides that dimerize into quantum dots, which serve as building blocks to further self-assemble into quantum confined supramolecular structures with diverse morphologies and photoluminescence properties. Especially, the emission can be tuned from the visible region to the near-infrared region (420 nm to 820 nm) by modulating the self-assembly process. Moreover, no obvious cytotoxic effect is observed for …

Biosynthetic Mechanism Of The Antibiotic Capuramycin, Erfu Yan

Theses and Dissertations--Pharmacy

A-102395 is a member of the capuramycin family of antibiotics which was isolated from the culture broth of Amycolatopsis sp. SANK 60206. A-102339 is structurally classified as a nucleoside antibiotic, which like all members of the capuramycin family, inhibits bacterial MraY (translocase I) with IC₅₀ of 11 nM which is the lowest among the capuramycin family. A semisynthetic derivative of capuramycin is currently in clinical trials as an antituberculosis antibiotic, suggesting high potential for using A-102395 as a starting point for new antibiotic discovery. In contrast to other capuramycins, A-102395 has a unique arylamine-containing polyamide side chain. The biosynthetic …

Optimisation Of Ion Exchange Chromatography Purification Protocols For A Staphylococcal Peptidoglycan Degrading Hydrolase Enzyme, Fiona Maher

Theses

Bacteriophage (phage) are the most abundant biological entities on earth and were first discovered by d’Herelle in 1917. They are found wherever their hosts live and, like all viruses they do not have the ability to make their own protein. Therefore, in order to reproduce, phage must invade and infect bacterial cells. This project focused on the optimisation of Ion Exchange Chromatography purification protocols for a staphylococcal peptidoglycan degrading hydrolase enzyme (CHAPk). The project objective was to obtain the greatest yield of enzyme from the growth of the E.coU XL 1-Blue expression system into which the vector pQE60 was previously …

Dielectric Properties Of Isolated Adrenal Chromaffin Cells Determined By Microfluidic Impedance Spectroscopy, A. C. Sabuncu, M. Stacey, G. L. Graviso, N. Semenova, P. T. Vernier, N. Leblanc, I. Chatterjee, J. Zaklit

Bioelectrics Publications

Knowledge of the dielectric properties of biological cells plays an important role in numerical models aimed at understanding how high intensity ultrashort nanosecond electric pulses affect the plasma membrane and the membranes of intracellular organelles. To this end, using electrical impedance spectroscopy, the dielectric properties of isolated, neuroendocrine adrenal chromaffin cells were obtained. Measured impedance data of the cell suspension, acquired between 1 kHz and 20 MHz, were fit into a combination of constant phase element and Cole-Cole models from which the effect of electrode polarization was extracted. The dielectric spectrum of each cell suspension was fit into a Maxwell-Wagner …

Quantitative Genetics Of Populus, Roba Bdeir

Dissertations, Master's Theses and Master's Reports

Evidence for bark, stem and stomatal density adaptation to different climates in the model species Populus is seen in both the natural population as well as in the greenhouse, but the genetic basis of these adaptation remains poorly understood. The present thesis investigates bark texture, bark thickness, diameter and stomatal density variations among Populus population using two quantitative genetics methods to attempt understand the genetic system controlling inheritance of these traits and to associate them with respective genes. The first approach aimed at detecting quantitative trait loci (QTL) associated with all phenotypic traits in an interspecific hybrid pedigree (Populus …

Manipulation Of Genes Involved In Secondary Cell Wall Development During Wood Formation In Poplar, Kavitha Satish Kumar

Dissertations, Master's Theses and Master's Reports

Lignins are second most abundant components of vascular plant cell walls. They provide plants with structural rigidity and are polymers of monolignols. Lignin polymerization is catalyzed by peroxidases and/or laccases. These enzymes are suggested to share functional overlap and mechanism by which they coordinate this process is not clearly understood. There are about 100 peroxidases and 50 laccase genes known in poplar genome out of which some stem differentiating xylem-specific (SDX) enzymes were selected for our study. The main objective was to genetically manipulate genes expressed in the SDX region in the cell wall to see the effects on lignin …

One-Step Hot Formamide Extraction Of Rna From Saccharomyces Cerevisiae, Daniel Shedlovskiy, Natalia Shcherbik, Dimitri G Pestov

Rowan-Virtua School of Osteopathic Medicine Departmental Research

Current methods for isolating RNA from budding yeast require lengthy and laborious steps such as freezing and heating with phenol, homogenization with glass beads, or enzymatic digestion of the cell wall. Here, extraction with a solution of formamide and EDTA was adapted to isolate RNA from whole yeast cells through a rapid and easily scalable procedure that does not require mechanical cell lysis, phenol, or enzymes. RNA extracted with formamide-EDTA can be directly loaded on gels for electrophoretic analysis without alcohol precipitation. A simplified protocol for downstream DNase treatment and reverse transcription reaction is also included. The formamide-EDTA extraction of …

High-Throughput Single-Molecule Telomere Characterization, Jennifer Mccaffrey, Eleanor Young, Katy Lassahn, Justin Sibert, Steven Pastor, Harold Riethman, Ming Xiao

School of Medical Diagnostics & Translational Sciences Publications

We have developed a novel method that enables global subtelomere and haplotype-resolved analysis of telomere lengths at the single-molecule level. An in vitro CRISPR/Cas9 RNA-directed nickase system directs the specific labeling of human (TTAGGG) n DNA tracts in genomes that have also been barcoded using a separate nickase enzyme that recognizes a 7bp motif genome-wide. High-throughput imaging and analysis of large DNA single molecules from genomes labeled in this fashion using a nanochannel array system permits mapping through subtelomere repeat element (SRE) regions to unique chromosomal DNA while simultaneously measuring the (TTAGGG) n tract length at the end of each …

Characterization Of Different Molecular Markers For Identification Of Salmonella Enterica Serovar Typhi In Pakistani Population, Faizan Muttiullah, Fida Muhammad Khan, Fakhar-I- Abbas, Sabiha Shamim

Journal of Bioresource Management

Typhoid is caused by Salmonella enterica serovar Typhi that is usually diagnosed by using serologic and immuno-chromatographic techniques in developing counties including Pakistan, which is thought to be an unreliable diagnostic method. For accurate diagnosis we used molecular techniques to amplify 204 bp StyR-36 and 498 bp flagellin gene for the identification of Salmonella enterica serovar Typhi. This study was done on 58 individuals diagnosed positive of typhoid via serologic tests and 50 healthy individuals as a control group. Success rate of amplification for flagellin gene was 77.58% while that for StyR-36 gene was 68.97% showing that flagellin gene primer …

Mutagenesis Of Human Alpha-Galactosidase A For The Treatment Of Fabry Disease, Erin Stokes

Dissertations, Theses, and Capstone Projects

Fabry disease is an X-linked lysosomal storage disorder caused by the deficiency of the enzyme, α-galactosidase A, which results in the accumulation of the lipid substrate. This accumulation results in obstruction of blood flow in patients and early demise at approximately 40-60 years of age. There is currently only one FDA approved treatment (Fabrazyme) classified as an enzyme replacement therapy. However, approximately 88% of patients experience a severe immune response that, rarely, can be fatal and is a huge cost burden at average $250,000 a year per patient. The structure of α-galactosidase A has been previously determined to be a …

Fret Biosensors: Engineering Fluorescent Proteins As Biological Tools For Studying Parkinson’S Disease, Nathan J. Leroy, Jacob R. Norley, Saranya Radhakrishnan, Mathew Tantama

The Summer Undergraduate Research Fellowship (SURF) Symposium

Parkinson’s Disease (PD) is a common neurodegenerative disease with over 200,000 new cases each year. In general, the cause of the disease is unknown, but oxidative stress inside of neurons has been associated with the disease’s pathology for some time. Currently, techniques to study the onset of PD inside of neurons are limited. This makes treatments and causes difficult to discover. One solution to this has been fluorescent protein biosensors. In short, these proteins can be engineered to glow when a certain state is achieved inside a cell. The present research discusses the engineering of a genetically-encoded fluorescent protein (FP) …

Fluorescent Protein Biosensor For Use In Parkinson's Research, Piper R. Miller, Keelan Trull, Mathew Tantama

The Summer Undergraduate Research Fellowship (SURF) Symposium

Purinergic signaling is a type of extracellular communication that occurs between cells, mediated by adenosine triphosphate (ATP), adenosine diphosphate (ADP), and adenosine. In Parkinson’s Disease, purinergic signaling is disrupted, which contributes to neurodegeneration. In order to monitor this change in cell-to-cell signaling, there is a need for the development of a fluorescent protein (FP) biosensor to study the changes in the concentration of the signaling molecule ATP and its decomposition bioproduct ADP. This summer a genetically encoded ADP sensor that measures changes in ADP concentration was developed. This sensor utilizes Forster Resonance Energy Transfer (FRET) which is a sensing technique …

Rational Drug Design Directed At Blocking The Initial Signaling Events In Lipopolysaccharide-Induced Sepsis., Christopher A. Tipton

Theses

Systemic Inflammatory Response Syndrome (SIRS) is classified as an immune system response to an infectious state. If left untreated, SIRS leads to sepsis, septic shock, end-organ dysfunction, and death. As a patient progresses through these stages, associations of acute respiratory distress, disseminated intravascular coagulation, and acute renal failure persist, resulting in millions of deaths annually. Lipopolysaccharide (LPS), a bacterial endotoxin, is released into the blood stream, triggering SIRS. LPS is found in the outer cell-wall of Gram-negative bacteria and is responsible for initiation of a devastating cytokine storm. One of the regions of LPS, lipid A, is a polyacylated glucosamine …