Biochemistry, Biophysics, and Structural Biology Commons^™

Analysis Of Bacterial Diversity In Arctic Soil And Sediment Samples From Ny-Alesund, Norway Using Denaturing Gradient Gel Electrophoresis., Yew Wen Chyin

Student Works (2010-2019)

Bacterial diversity in soil and sediment samples from eighteen sites in and around Dasan Station, Ny-Ålesund, Norway was analyzed using Denaturing Gradient Gel Electrophoresis (DGGE), and was correlated with the environmental variables. Samples analysed included terrestrial soils, beach soil, periglacier soils, inland lake bank sediments, melt lake sediments, and marine sediments. Spearman rank order correlation showed significant correlations (P = 0.001) between conductivity (r = 0.470) and pH (r = 0.294) with the bacterial diversity. Soil from the tundra site (sample 17) appeared to have the most diverse bacterial community while inland lake bank sediment from the …

Frustrated Drift Of An Anchored Scroll-Wave Filament And The Geodesic Principle, Marcel Wellner, Christian W. Zemlin, Arkady M. Pertsov

Bioelectrics Publications

We investigate anchored scroll-wave filaments in an excitable medium whose diffusivity matrix, including its determinant, is spatially nonuniform. The study is motivated by cardiological applications where scroll-wave behavior in the presence of diffusivity gradients is believed to play an important role in the development of severe arrhythmias. A diffusivity gradient is expected to make the filament drift, unless drift is prevented ("frustrated") by anchoring to localized defects in the propagation medium. The resulting stationary filament is a geodesic curve, as demonstrated here in the case of a nonzero but constant gradient. That is, the diffusivity matrix has a determinant that …

Electroporation-Mediated Delivery Of A Naked Dna Plasmid Expressing Vegf To The Porcine Heart Enhances Protein Expression, W. G. Marshall Jr., B. A. Boone, J. D. Burgos, S. I. Gografe, M. K. Baldwin, M. L. Danielson, M. J. Larson, D. R. Caretto, Y. Cruz, B. Ferraro, L. C. Heller, K. E. Ugen, M. J. Jaroszeski, R. Heller

Bioelectrics Publications

Gene therapy is an attractive method for the treatment of cardiovascular disease. However, using current strategies, induction of gene expression at therapeutic levels is often inefficient. In this study, we show a novel electroporation (EP) method to enhance the delivery of a plasmid expressing an angiogenic growth factor (vascular endothelial growth factor, VEGF), which is a molecule previously documented to stimulate revascularization in coronary artery disease. DNA expression plasmids were delivered in vivo to the porcine heart with or without coadministered EP to determine the potential effect of electrically mediated delivery. The results showed that plasmid delivery through EP significantly …

Mzm1 Influences A Labile Pool Of Mitochondrial Zinc Important For Respiratory Function, Aaron Atkinson, Oleh Khalimonchuk, Pamela Smith, Hana Sabic, David Eide, Dennis R. Winge

Department of Biochemistry: Faculty Publications

Zinc is essential for function of mitochondria as a cofactor for

several matrix zinc metalloproteins. We demonstrate that a

labile cationic zinc component of low molecular mass exists in

the yeast mitochondrial matrix. This zinc pool is homeostatically

regulated in response to the cellular zinc status. This pool

of zinc is functionally important because matrix targeting of a

cytosolic zinc-binding protein reduces the level of labile zinc

and interferes with mitochondrial respiratory function. We

identified a series of proteins that modulate the matrix zinc

pool, one of which is a novel conserved mitochondrial protein

designated Mzm1. Mutant mzm1∆ cells …

Identification And Characterization Of Small Compound Inhibitors Of Human Fatp2, Angel Sandoval, Aalap Chokshi, Elliot D. Jesch, Paul N. Black, Concetta C. Dirusso

Department of Biochemistry: Faculty Publications

Fatty acid transport proteins (FATPs) are bifunctional proteins, which transport long chain fatty acids into cells and activate very long chain fatty acids by esterification with coenzyme A. In an effort to understand the linkage between cellular fatty acid transport and the pathology associated with excessive accumulation of exogenous fatty acids, we targeted FATP-mediated fatty acid transport in a high throughput screen of more than 100,000 small diverse chemical compounds in yeast expressing human FATP2 (hsFATP2). Compounds were selected for their ability to depress the transport of the fluorescent long chain fatty acid analogue, C₁-BODIPY-C₁₂. Among …

Uncultivated Environmental Tm7 Model To Study Human Disease-Associated Tm7 Bacteria, David Barton

Master's Theses

The TM7 bacterial phylum has no cultivated species and includes members that span a broad range of environmental and human habitats, some of which are associated with human periodontitis. In this project, activated wastewater TM7 bacteria were analyzed and their relatedness compared to human-associated TM7 bacteria for the long-term goal of using an environmental TM7 to better understand TM7 pathogenesis in humans. DNA was extracted from activated wastewater and PCR amplified using TM7 16S rRNA gene- specific primers. The ~1,170 base pair PCR products were then cloned and sequenced. DNA sequencing and phylogenetic analysis identified environmental TM7 clones with high …

Evolution Of New Enzymatic Function By Structural Modulation Of Cysteine Reactivity In Pseudomonas Fluorescens Isocyanide Hydratase, Mahadevan Lakshminarasimhan, Peter Madzelan, Ruth Nan, Nicole Marie Milkovic, Mark A. Wilson

Department of Biochemistry: Faculty Publications

Isocyanide (formerly isonitrile) hydratase (EC 4.2.1.103) is an enzyme of the DJ-1 superfamily that hydrates isocyanides to yield the corresponding N-formamide. In order to understand the structural basis for isocyanide hydratase (ICH) catalysis, we determined the crystal structures of wild-type and several site-directed mutants of Pseudomonas fluorescens ICH at resolutions ranging from 1.0 to 1.9 Å. We also developed a simple UV-visible spectrophotometric assay for ICH activity using 2-naphthyl isocyanide as a substrate. ICH contains a highly conserved cysteine residue (Cys101) that is required for catalysis and interacts with Asp17, Thr102, and an ordered water molecule in the active …

Identification Of Deletion Mutants Of Inositol Kinases And Phosphatases Hypersensitive To Valproate, W-Mudiyanselage Manoj Senaka Bandara

Wayne State University Theses

ABSTRACT

IDENTIFICATION OF DELETION MUTANTS OF INOSITOL KINASES AND PHOSPHATASES HYPERSENSITIVE TO VALPROATE

by

WELLEWATTA MUDIYANSELAGE MANOJ SENAKA BANDARA

MAY 2010

Advisor: Dr. Miriam Greenberg

Major: Biological Sciences

Degree: Master of Science

Bipolar disorder (BD) is a chronic psychiatric illness affecting at least 1% of the world population. BD is ranked as the sixth greatest cause of death or disability globally. The cause of BD is unknown. Although the anticonvulsant valproate (VPA) is widely used as a mood stabilizer to treat BD, VPA is not completely effective and causes numerous side effects. Hence, it is important to develop more effective …

The Roles Of Rhle And Hfq In Srna-Dependent Gene Regulation, Abeykoon Jayalath Iresha Sandeepanie Rathnayake

Wayne State University Theses

THE ROLES OF RHLE AND HFQ IN SRNA-DEPENDENT GENE REGULATION

by

ABEYKOON JAYALATH IRESHA SANDEEPANIE RATHNAYAKE

DECEMBER 2010

Advisor: Dr. Andrew Feig

Major: Chemistry

Degree: Master of Science

Bacteria are adapted to live in diverse environmental conditions. Thus, they show excellent tolerance and response to extreme environmental conditions caused by low or high temperatures, high salinity, reactive oxygen species or high nutrient concentrations. sRNAs have been identified and characterized as cis-acting or trans-acting post-transcriptional regulators in diverse cellular processes including virulence and adaptation to environmental stress (11,12,14,15). Interactions of sRNAs and target mRNAs result in translational repression, translational activation or/and …

Creg1 And Its Enhancement Of P16ink4a-Induced Senescence, Benchamart Moolmuang

Wayne State University Dissertations

Bypassing cellular senescence, an irreversible growth arrest of cells that is activated in normal cells to become immortal is one of the prerequisites for carcinogenesis. Cellular senescence can be triggered by shortening of telomeres and certain cellular stresses. Using spontaneously immortalized Li-Fraumeni Syndrome (LFS) fibroblasts, we found that CREG1 (Cellular Repressor of E1A-stimulated Genes1) is one of genes whose expression fit the criteria of senescence-associated genes, decreased expression during immortalization and increased in senescence. Moreover, we found that epigenetic mechanisms regulate CREG1 expression in LFS fibroblasts. CREG1 is a secreted glycoprotein that was shown to bind Rb-family pocket proteins and …

Group Ii Intron Dynamics In Heterologous Hosts, Venkata Raghavendra Aditya Chalamcharla

Legacy Theses & Dissertations (2009 - 2024)

Group II introns are ribozymes with an innate ability to self-splice. They are found predominantly in bacterial and bacterial-derived organellar genomes, but not in the nuclear genomes of eukaryotes. In bacteria, group II introns often behave as mobile retroelements, invading host DNA and exploiting its machinery to complete the retromobility process. The object of my studies is the group II intron found in the Lactococcus lactis relaxase gene. To determine the nature of the group II intron-host relationship, we performed a genetic screen and identified several host factors that affect group II intron retromobility in Escherichia coli, which provides a …

Preparation And Application Of Transgenic And Knockout Mouse Models For Studies On Cytochrome P450 Functions, Yuan Wei

Legacy Theses & Dissertations (2009 - 2024)

The overall objective of this dissertation is to study the in vivo function of microsomal cytochrome P450 monooxygenases (P450s), which metabolize numerous drugs, chemical carcinogens, environmental pollutants, as well as endogenous signaling molecules such as steroid hormones and eicosanoids. The major research tool of this study involves the development of transgenic and knockout mouse models. The specific aims are 1) to study the in vivo function of NADPH-cytochrome P450 reductase (CPR) and CPR-dependent enzymes using a mouse model with a reversible hypomorphic Cpr gene; 2) to study the in vivo function of CYP2A13 with a CYP2A13- transgenic model; and 3) …

The Alliinase And Lachrymatory Factor Synthase Systems In Petiveria Alliacea, Quan He

Legacy Theses & Dissertations (2009 - 2024)

Isolation and characterization of an alliinase and a lachrymatory factor synthase from the Amazonian medicinal plant Petiveria alliacea is reported. The alliinase is unusual in that it is heteromeric, whereas all previous reports of characterized alliinases have shown them to be monomers or homomultimers. The protein possesses 5 subunits: two alpha subunits which are glycosylated and connected by a disulfide bond, and beta, gamma and delta subunits. The alliinase exhibits broad substrate specificity, reacting with S-substituted-L-cysteine sulfoxides with aromatic, aliphatic, alkenyl and polar side chains. The alliinase-mediated breakdown of a variety of cysteine sulfoxide derivatives yields transitory sulfenic acid intermediate …

Saturation Transfer Difference Nmr Studies : Pyrazinamide And Pyrazinamide Analogs Coordinating With Mycobacterium Tuberculosis Fatty Acid Synthase I (Fas I), Halimah Sayahi

Legacy Theses & Dissertations (2009 - 2024)

One third of the world's population is infected with M. tuberculosis, the causative agent of tuberculosis (TB) that claims the life of approximately 2 million people every year. The rapid spread of multidrug-resistance tuberculosis as well the emergence of extensively drug resistance strains that are resistant to virtually any known antibiotic, has transformed this once curable disease into a major public health challenge. To regain control of this disease and the associated mortality and morbidity new antibiotics are urgently needed.

Elucidating The Structure Of Protein Aggregates By Raman Spectroscopy, Ludmila A. Popova

Legacy Theses & Dissertations (2009 - 2024)

The structures and properties of amyloid fibrils are of considerable interest due to their associations with numerous neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease and transmissible spongiform encephalopaties (prion diseases). Understanding fibrillogenesis at a molecular level requires detailed structural characterization of amyloid fibrils. However amyloid fibrils are difficult objects to study due to their non-crystalline and insoluble nature. These properties make the application of classical tools of structural biology, such as X-Ray crystallography and solution Nuclear Magnetic Resonance spectroscopy, impractical for structural characterization of protein fibrils.

Morphological Changes And Immunohistochemical Expression Of Rage And Its Ligands In The Sciatic Nerve Of Hyperglycemic Pig (Sus Scrofa), Judyta K. Juranek, Alexey Aleshin, Eileen M. Rattigan, Lynne Johnson, Wu Qu, Fei Song, Radha Ananthakrishnan, Nosirudeen Quadri, Shi Du Yan, Ravichandran Ramasamy, Ann Marie Schmidt, Matthew S. Geddis

Publications and Research

The aim of our project was to study the effect of streptozotocin (STZ)—induced hyperglycemia on sciatic nerve morphology, blood plasma markers and immunohistochemical expression of RAGE (the Receptor for Advanced Glycation End-products), and its ligands—S100B and Carboxymethyl Lysine (CML)-advanced glycation endproduct (AGE) in the laboratory pig. Six months after STZ—injections, blood plasma measurements, morphometric analysis of sciatic nerve fiber density, immunofluorescent distribution of potential molecular neuropathy contributors, ELISA measurement of plasma AGE level and HPLC analysis of sciatic nerve levels of one of the pre-AGE and the glycolysis intermediate products—methyl-glyoxal (MG) were performed. The results of our study revealed that …

Patterns Of Apoptotic Poly(Adp-Ribose) Polymerase Cleavage Induced By Laromustine And Its Analogs, Adam N. Paine

Honors Theses

The anticancer prodrug Laromustine (VNP40101M) has produced promising remission rates in clinical trials among leukemic patients relative to currently available chemotherapeutics. Such improvements demand that the agent’s mechanism of action be elucidated. This study aimed to determine the role of poly(ADP-ribose) polymerase (PARP) in Laromustine-induced cell death. Previous studies indicated that the methyl isocyanate produced upon activation of Laromustine is largely responsible for its effective induction of apoptosis. Results reported herein strongly support the activation of a PARP-dependent apoptotic pathway by Laromustine’s carbamoylating and chlorethylating subspecies. Furthermore, it is evident that the Laromustine-induced PARP-dependent apoptosis is primarily attributable to methyl …

Investigating The Metal Binding Sites In Znta, A Zinc Transporting Atpase, Sandhya Muralidharan

Wayne State University Dissertations

ZntA from Escherichia coli is a member of the P_IBtype ATPase family of transporters. The P_IB-type ATPase pumps maintain cellular homeostasis of heavy metals such as Zn²⁺, Co²⁺, Cu²⁺, Cu⁺, and mediate resistance to toxic metals such as Pb²⁺, Cd²⁺ and Ag⁺. ZntA confers resistance to Pb²⁺, Cd²⁺, and Zn²⁺ by pumping these ions out of the cytoplasm. ZntA has two metal binding sites, one in the hydrophilic N-terminal domain and the other in the transmembrane region. The …

Towards An Understanding Of The Etiology Of Abdominal Aortic Aneurysms: Identification Of Genes Implicated In Aaa Risk And Development, John Hunt Lillvis

Wayne State University Dissertations

Abdominal aortic aneurysm (AAA) is a common disease for which mechanisms of formation are still not well understood. Despite a strong genetic component to AAA risk, specific risk alleles are still largely unidentified. AAA is also a localized disease with a majority occurring in the infrarenal abdominal aorta and is six times more common than aneurysms of the thoracic aorta. To determine whether risk alleles are present in functional positional candidate genes. we: 1. performed a genetic association study using DNA from AAA cases and controls in ten candidate genes and 2. performed exon sequencing on three genes with evidence …

Interactions Of The Cellular Sumoylation System With Influenza A Virus And Its Non-Structural Protein Ns1a (Ns1a), Sangita Pal

Open Access Theses & Dissertations

The most important current anti-influenza weapons, vaccination and antiviral drugs, can be rapidly rendered fully ineffective thanks to the virus's high mutational rate, which produces viruses exhibiting new antigenic properties and structural proteins insensitive to the drug's mechanism of action. One attractive alternative is to develop drugs that modulate the activity of cellular systems either required for viral growth or able to neutralize viral growth. Here we demonstrate that the cellular SUMOylation system, a post-translational modification involving the conjugation of the Small Ubiquitin-like MOdifier (SUMO) to specific protein targets using a Ubiquitin-like enzymatic cascade, interacts closely with influenza virus during …