Molecular Biology Commons^™

Genetically Engineering Of Synechocystis Pcc 6803 For Sustainable Production Of Cinnamic Acid, Shahrah Saad Alqahtani

Theses and Dissertations

Cinnamic acid is the precursor for phenylpropanoids, which play important roles in plant growth, development, and stress responses. It is highly beneficial to human health for its anticancer, antitoxic and anti-inflammatory functions. Currently, the major methods of industrial production of cinnamic acid are chemical synthesis and extraction from plants. However, these methods are often associated with low production yields and high costs. Cyanobacterial platforms could be highly suitable for phenylpropanoid production due to potential high yields and eco- friendly, cost- effective production processes. The primary purpose of this study is to explore the possibility of using the cyanobacterium Synechocystis PCC …

C. Elegans Avoids Toxin-Producing Streptomyces Using A Seven Transmembrane Domain Chemosensory Receptor, Alan Tran, Angelina Tang, Colleen O’Loughlin, Anthony Balistreri, Eric Chang, Doris Coto Villa, Joy Li, Aruna Varshney, Vanessa Jimenez, Jacqueline Pyle, Bryan Tsujimoto, Christopher Wellbrook, Christopher Vargas, Alex Duong, Nebat Ali, Sarah Matthews, Samantha Levinson, Sarah Woldemariam, Sami Khuri, Martina Bremer, Daryl Eggers, Noelle L’Etoile, Laura Miller Conrad, Miri Vanhoven

Faculty Publications, Chemistry

Predators and prey co-evolve, each maximizing their own fitness, but the effects of predator–prey interactions on cellular and molecular machinery are poorly understood. Here, we study this process using the predator Caenorhabditis elegans and the bacterial prey Streptomyces, which have evolved a powerful defense: the production of nematicides. We demonstrate that upon exposure to Streptomyces at their head or tail, nematodes display an escape response that is mediated by bacterially produced cues. Avoidance requires a predicted G-protein-coupled receptor, SRB-6, which is expressed in five types of amphid and phasmid chemosensory neurons. We establish that species of Streptomyces secrete dodecanoic acid, …

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 …

A Combined Computational Strategy Of Sequence And Structural Analysis Predicts The Existence Of A Functional Eicosanoid Pathway In Drosophila Melanogaster, Michael Scarpati

Dissertations, Theses, and Capstone Projects

With increased understanding of their roles in signal transduction and metabolism, eicosanoids have emerged as important players in human health and disease. Mammalian prostanoids and related lipid mediators perform varied functions in different tissues and organs. Synthesized through the oxygenation of C20 polyunsaturated fatty acids, mammalian eicosanoids are both pro- and anti-inflammatory. The physiological contexts in which eicosanoid family members act at the cellular level are not well understood. In this study, we examined whether the genome of Drosophila melanogaster, a powerful model for innate immunity and inflammation, codes for the enzymes required for eicosanoid biosynthesis. We report the …

Insight Into The Interaction Between The Peroxisome Proliferator-Activated Receptor Gamma (Pparγ) And Adipocyte Fatty Acid-Binding Protein (A-Fabp), Qian Wang

Dissertations, Theses, and Capstone Projects

The Adipocyte Fatty Acid-Binding Protein (AFABP) is mainly expressed in fat cells. It can bind fatty acids and other lipophilic substances such as eicosanoids and retinoids. The peroxisome proliferator-activated receptor γ (PPARγ) is a nuclear receptor protein that requires ligand binding to regulate the specific gene transcription. PPARγ is expressed at extremely high levels in adipose tissue, macrophages, and the large intestine, where it controls lipid adipogenesis and energy conversion. Moreover, it has been found that AFABP and PPARγ can form a complex in vivo. It is proposed that AFABP carries the ligand and enters into the nucleus where it …

Engineered Aptamers To Probe Molecular Interactions On The Cell Surface, Sana Batool, Sanam Bhandari, Shanell George, Precious Okeoma, Nabeela Van, Hazan E. Zümrüt, Prabodhika Mallikaratchy

Publications and Research

Significant progress has been made in understanding the nature of molecular interactions on the cell membrane. To decipher such interactions, molecular scaffolds can be engineered as a tool to modulate these events as they occur on the cell membrane. To guarantee reliability, scaffolds that function as modulators of cell membrane events must be coupled to a targeting moiety with superior chemical versatility. In this regard, nucleic acid aptamers are a suitable class of targeting moieties. Aptamers are inherently chemical in nature, allowing extensive site-specific chemical modification to engineer sensing molecules. Aptamers can be easily selected using a simple laboratory-based in …

Differential Uptake Of Gold Nanoparticles By 2 Species Of Tadpole, The Wood Frog (Lithobates Sylvaticus) And The Bullfrog (Lithobates Catesbeianus), Lucas B. Thompson, Gerardo L.F. Carfagno, Kurt Andresen, Andrea J. Sitton, Taylor B. Bury, Laura L. Lee, Kevin T. Lerner, Peter P. Fong

Biology Faculty Publications

Engineered nanoparticles are aquatic contaminants of emerging concern that exert ecotoxicological effects on a wide variety of organisms. We exposed cetyltrimethylammonium bromide–capped spherical gold nanoparticles to wood frog and bullfrog tadpoles with conspecifics and in combination with the other species continuously for 21 d, then measured uptake and localization of gold. Wood frog tadpoles alone and in combination with bullfrog tadpoles took up significantly more gold than bullfrogs. Bullfrog tadpoles in combination with wood frogs took up significantly more gold than controls. The rank order of weight-normalized gold uptake was wood frogs in combination > wood frogs alone > bullfrogs in combination …

Translational Fidelity, Mistranslation, And The Cellular Responses To Stress, Kyle Mohler, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Faithful translation of mRNA into the corresponding polypeptide is a complex multistep process, requiring accurate amino acid selection, transfer RNA (tRNA) charging and mRNA decoding on the ribosome. Key players in this process are aminoacyl-tRNA synthetases (aaRSs), which not only catalyse the attachment of cognate amino acids to their respective tRNAs, but also selectively hydrolyse incorrectly activated non-cognate amino acids and/or misaminoacylated tRNAs. This aaRS proofreading provides quality control checkpoints that exclude non-cognate amino acids during translation, and in so doing helps to prevent the formation of an aberrant proteome. However, despite the intrinsic need for high accuracy during translation, …

Itraq-Based Proteomics Analysis And Network Integration For Kernel Tissue Development In Maize, Long Zhang, Yongbin Dong, Qilei Wang, Chunguang Du, Wenwei Xiong, Xinyu Li, Sailan Zhu, Yuling Li

Department of Biology Faculty Scholarship and Creative Works

Grain weight is one of the most important yield components and a developmentally complex structure comprised of two major compartments (endosperm and pericarp) in maize (Zea mays L.), however, very little is known concerning the coordinated accumulation of the numerous proteins involved. Herein, we used isobaric tags for relative and absolute quantitation (iTRAQ)-based comparative proteomic method to analyze the characteristics of dynamic proteomics for endosperm and pericarp during grain development. Totally, 9539 proteins were identified for both components at four development stages, among which 1401 proteins were non-redundant, 232 proteins were specific in pericarp and 153 proteins were specific in …

Molecular Mechanisms Of C-Terminal Eps15 Homology Domain Containing (Ehd) Protein Function, Kriti Bahl

Theses & Dissertations

Endocytic trafficking is not only an essential process for the maintenance of cellular homeostasis but also plays a vital role in regulating diverse cellular processes such as signaling, migration and cell division. The C-terminal Eps 15 Homology Domain proteins (EHD1-4) play pivotal roles in regulating distinct steps of endocytic trafficking. Among the EHDs, EHD2 is disparate both in terms of sequence homology (70%) and its subcellular localization at the caveolae. The crystal structure of EHD2 has been solved and it contains an unstructured loop consisting of two proline-phenylalanine (PF) motifs: KPFRKLNPF. However, the other paralogs EHD1, …

Engaging Medical Students In Problem-Based Search And Study Of The Biomedical Literature., Zhiyong Han, Samantha L Margulies, Divya Kurian, Joshua M Jabaut, Akshita Mehta, Ramzi Dudum, Huberta Koudoro, Ria S Roberts, Jay Lee, Jonathan Li, Hieu T Nguyen, Mark Elliott

Biochemistry and Molecular Medicine Faculty Publications

No abstract provided.

Thermodynamic And Kinetic Analyses Of Iron Response Element (Ire)-Mrna Binding To Iron Regulatory Protein, Irp1, Mateen A. Khan, William E. Walden, Elizabeth C. Theil, Dixie J. Goss

Publications and Research

Comparison of kinetic and thermodynamic properties of IRP1 (iron regulatory protein1) binding to FRT (ferritin) and ACO2 (aconitase2) IRE-RNAs, with or without Mn2+, revealed differences specific to each IRE-RNA. Conserved among animal mRNAs, IRE-RNA structures are noncoding and bind Fe2+ to regulate biosynthesis rates of the encoded, iron homeostatic proteins. IRP1 protein binds IRERNA, inhibiting mRNA activity; Fe2+ decreases IRE-mRNA/IRP1 binding, increasing encoded protein synthesis. Here, we observed heat, 5 °C to 30 °C, increased IRP1 binding to IRE-RNA 4-fold (FRT IRE-RNA) or 3-fold (ACO2 IRE-RNA), which was enthalpy driven and entropy favorable. Mn2+ (50 μM, 25 °C) increased IRE-RNA/IRP1 …

Dietary Phytochemicals As Inhibitors Of Primary Amine Oxidase, Padraig Shanahan, Jeffrey O'Sullivan, Keith F. Tipton, Gemma Kinsella, Barry J. Ryan, Gary T. Henehan

Articles

Phytochemicals such as methylxanthines, catechins and polyphenols show health benefits in a range of diseases although their mechanism of action is not fully understood. Primary Amine Oxidase (PrAO) is widely recognised as a therapeutic drug target for the treatment of inflammatory, vascular and neurodegenerative diseases. Previous work in our laboratories showed that caffeine inhibited bovine PrAO activity with a Ki of 1.0mM. In the present study we examined a range of methylxanthines and catechins as inhibitors of bovine PrAO. The methylxanthines tested were caffeine, paraxanthine, theophylline, theobromine and 7-methylxanthine. Of these, only theobromine was an inhibitor with an IC50 of …

Biophysical And Computational Studies Of The Vcci:Vmip-Ii Complex, Anna Nguyen, Nai-Wei Kuo, Laura Showalter, Ricardo Ramos, Cynthia Dupureur, Michael Colvin, Patricia Liwang

Chemistry & Biochemistry Faculty Works

Certain viruses have the ability to subvert the mammalian immune response, including interference in the chemokine system. Poxviruses produce the chemokine binding protein vCCI (viral CC chemokine inhibitor; also called 35K), which tightly binds to CC chemokines. To facilitate the study of vCCI, we first provide a protocol to produce folded vCCI from Escherichia coli (E. coli.) It is shown here that vCCI binds with unusually high affinity to viral Macrophage Inflammatory Protein-II (vMIP-II), a chemokine analog produced by the virus, human herpesvirus 8 (HHV-8). Fluorescence anisotropy was used to investigate the vCCI:vMIP-II complex and shows that vCCI binds to …

Metastasis-Associated Protein 1 Deficiency Results In Compromised Pulmonary Alveolar Capillary Angiogenesis In Mice., Jun-Hui Qin, Zhen-Yu Ke, Qiang Zhou, Li Wang, Yuan Liang, Ying-Mei Wang, Tong Yang, Xing Gao, Jing Ye, Rekesh Kumar, Rui-An Wang

Biochemistry and Molecular Medicine Faculty Publications

Background

The aim of this study was to investigate the effects of metastasis-associated protein 1 (MTA1) deficiency during angiogenesis of pulmonary alveolar capillaries in mice and to determine the molecular mechanisms involved.

Material/Methods

The expressions of MTA1, CD34, vascular endothelial growth factor (VEGF), alpha smooth muscle actin (α-SMA), and HIF-1α were analyzed in the lungs of MTA1-knockout (KO) and wild-type mice at embryonic day 18.5 and 2 months by quantitative PCR, immunoblotting, and immunohistochemistry. The morphological changes were investigated during pulmonary alveolar capillary formation. The heart weight/body weight (HW/BW) ratio and the size of the right ventricular wall cardiomyocytes …

Carbonyl Reduction By Ymfi Completes The Modification Of Ef-P In Bacillus Subtilis To Prevent Accumulation Of An Inhibitory Modification State, Katherine R. Hummels, Anne Witzky, Andrei Rajkovic, Rodney Tollerson Ii, Lisa A. Jones, Michael Ibba, Daniel B. Kearns

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Translation elongation factor P (EF‐P) in Bacillus subtilis is required for a form of surface migration called swarming motility. Furthermore, B. subtilis EF‐P is post‐translationally modified with a 5‐aminopentanol group but the pathway necessary for the synthesis and ligation of the modification is unknown. Here we determine that the protein YmfI catalyzes the reduction of EF‐P‐5 aminopentanone to EF‐P‐5 aminopentanol. In the absence of YmfI, accumulation of 5‐aminopentanonated EF‐P is inhibitory to swarming motility. Suppressor mutations that enhanced swarming in the absence of YmfI were found at two positions on EF‐P, including one that changed the conserved modification site (Lys …

Performing A Genetic Screen To Identify Factors That Promote Lncrna-Dependent Gene Repression, Chrishan Fernando, Cecilia Yiu, Sara Cloutier, Siwen Wang, Elizabeth Tran

The Summer Undergraduate Research Fellowship (SURF) Symposium

Long non-coding RNAs (lncRNAs) were once thought not to have useful functions in organisms but rather to be products of aberrant transcription. However, roles are being found for lncRNAs in beneficial processes such as controlling gene expression. In some of these cases, lncRNAs form R-loops in vivo. R-loops are nucleic acid structures consisting of hybridized strands of single-stranded DNA (ssDNA) and single-stranded RNA (ssRNA) as well as the displaced strand of ssDNA. Formation of these R-loops is important for gene regulation by the lncRNAs. However, factors that promote formation of lncRNA R-loops are not known. The gene PHO84 is being …

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 …

Bow-Tie Signaling In C-Di-Gmp: Machine Learning In A Simple Biochemical Network, Jinyuan Yan, Maxime Deforet, Kerry E. Boyle, Rayees Rahman, Raymond Liang, Chinweike Okegbe, Lars E. P. Dietrich, Weigang Qiu, Joao B. Xavier

Publications and Research

Bacteria of many species rely on a simple molecule, the intracellular secondary messenger c-di-GMP (Bis-(3'-5')-cyclic dimeric guanosine monophosphate), to make a vital choice: whether to stay in one place and form a biofilm, or to leave it in search of better conditions. The c-di-GMP network has a bow-tie shaped architecture that integrates many signals from the outside worldÐthe input stimuliÐinto intracellular c-di-GMP levels that then regulate genes for biofilm formation or for swarming motilityÐthe output phenotypes. How does the `uninformed' process of evolution produce a network with the right input/output association and enable bacteria to make the right choice? Inspired …