Biochemistry, Biophysics, and Structural Biology Commons^™

Inflammasome-Dependent Caspase-1 Activation In Cervical Epithelial Cells Stimulates Growth Of The Intracellular Pathogen Chlamydia Trachomatis, Ali A. Abdul-Sater, Evonne Koo, Georg Hacker, David M. Ojcius

All Dugoni School of Dentistry Faculty Articles

Inflammasomes have been extensively characterized in monocytes and macrophages, but not in epithelial cells, which are the preferred host cells for many pathogens. Here we show that cervical epithelial cells express a functional inflammasome. Infection of the cells by Chlamydia trachomatis leads to activation of caspase-1, through a process requiring the NOD-like receptor family member NLRP3 and the inflammasome adaptor protein ASC. Secretion of newly synthesized virulence proteins from the chlamydial vacuole through a type III secretion apparatus results in efflux of K+ through glibenclamide-sensitive K+ channels, which in turn stimulates production of reactive oxygen species. Elevated levels of reactive …

Gas Phase Chiral Recognition, Characterization Of Porous Polymer Monolith Nanospray Ionization, And The Negative Mode Crafti Method Using Fourier Transform Ion Cyclotron Resonance Mass Spectrometry, Nannan Fang

Theses and Dissertations

Our group has been studying chiral recognition in gas phase using mass spectrometry for more than 10 years. We are interested in gas phase studies of fundamental interactions because the gas phase avoids complications and masking effects that may arise upon solvation. Therefore, the results of gas phase experiments can be directly compared with those of high-level computational studies. In chapter 2, I studied the roles of hydrogen bonding and pi stacking in gas phase chiral recognition between aromatic crown molecules and aromatic amines. High affinity between host and guest doesn't necessarily result in better recognition. If the affinity is …

Broad Range Amino Acid Specificity Of Rna-Dependent Lipid Remodelling By Multiple Peptide Resistance Factors, Hervé Roy, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Aminoacylphosphatidylglycerol synthases (aaPGSs) are multiple peptide resistance factors that transfer amino acids from aminoacyl-tRNAs to phosphatidylglycerol (PG) in the cytoplasmic membrane. Aminoacylation of PG is used by bacteria to decrease the net negative charge of the cell envelope, diminishing affinity for charged molecules and allowing for adaptation to environmental changes. Lys-PGS, which transfers lysine to PG, is essential for the virulence of certain pathogens, providing resistance to both host cationic antimicrobial peptides and therapeutic antibiotics. Ala-PGS was also recently described, but little is known about the possible activities of other members of the highly diverse aaPGS family of proteins. Systematic …

Self-Assembly And Disassembly Of The Snare Complex: Examined Using Circular Dichroism And Atomic Force Microscopy, Jeremy D. Cook, Won Jin Cho, Timothy L. Stemmler, Bhanu P. Jena

Biochemistry and Molecular Biology Faculty Publications

In this study, we report for the first time that both t-SNAREs and v-SNARE and their complexes in buffered suspension, exhibit defined peaks at CD signals of 208 and 222 nm wavelengths, consistent with a higher degree of helical secondary structure. Surprisingly, when incorporated in lipid membrane, both SNAREs and their complexes exhibit reduced folding. In presence of NSF-ATP, the SNARE complex disassembles, as reflected from the CD signals demonstrating elimination of α-helices within the structure.

The Cca Anticodon Specifies Separate Functions Inside And Outside Translation In Bacillus Cereus, Sandro F. Ataide, Theresa E. Rogers, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Bacillus cereus 14579 encodes two tRNAs with the CCA anticodon, tRNA^Trp and tRNA^Other. tRNA^Trp was separately aminoacylated by two enzymes, TrpRS1 and TrpRS2, which share only 34% similarity and display different catalytic capacities and specificities. TrpRS1 was 18-fold more proficient at aminoacylating tRNA^Trp with Trp, while TrpRS2 more efficiently utilizes the Trp analog 5-hydroxy Trp. tRNA^Other was not aminoacylated by either TrpRS but instead by the combined activity of LysRS1 and LysRS2, which recognized sequence elements absent from tRNA^Trp. Polysomes were found to contain tRNA^Trp, consistent with its role in …

Microbial Nad Metabolism: Lessons From Comparative Genomics, Francesca Gazzaniga, Rebecca Stebbins, Sheila Z. Chang, Mark A. Mcpeek, Charles Brenner

Dartmouth Scholarship

NAD is a coenzyme for redox reactions and a substrate of NAD-consuming enzymes, including ADP-ribose transferases, Sir2-related protein lysine deacetylases, and bacterial DNA ligases. Microorganisms that synthesize NAD from as few as one to as many as five of the six identified biosynthetic precursors have been identified. De novo NAD synthesis from aspartate or tryptophan is neither universal nor strictly aerobic. Salvage NAD synthesis from nicotinamide, nicotinic acid, nicotinamide riboside, and nicotinic acid riboside occurs via modules of different genes. Nicotinamide salvage genes nadV and pncA, found in distinct bacteria, appear to have spread throughout the tree of life …

Theoretical Studies Of 2,3 -Sigmatropic Rearrangements Of Allylic Selenoxides And Selenimides, Craig A. Bayse, Sonia Antony

Chemistry & Biochemistry Faculty Publications

Density-functional theory is used to model the endo and exo transition states for [2,3]-sigmatropic rearrangement of allylic aryl-selenoxides and -selenimides. The endo transition state is generally preferred for selenoxides if there is no substitution at the 2 position of the allyl group. Based upon the relative energies of the endo and exo transition states, enantioselectivity of rearrangements is expected to be greatest for molecules with substitutions at the 1- or (E)-3- position of the allyl group. Ortho substitution of a nitro group on the ancillary selenoxide phenyl ring reduces the activation barriers, increases the difference between the endo and exo …

The Crystal Structure Of The Escherichia Coli Autoinducer-2 Processing Protein Lsrf, Zamia Caridad Diaz , '09, K. B. Xavier, Stephen T. Miller

Chemistry & Biochemistry Faculty Works

Many bacteria produce and respond to the quorum sensing signal autoinducer-2 (AI-2). Escherichia coli and Salmonella typhimurium are among the species with the lsr operon, an operon containing AI-2 transport and processing genes that are up regulated in response to AI-2. One of the Lsr proteins, LsrF, has been implicated in processing the phosphorylated form of AI-2. Here, we present the structure of LsrF, unliganded and in complex with two phospho-AI-2 analogues, ribose-5-phosphate and ribulose-5-phosphate. The crystal structure shows that LsrF is a decamer of (alpha beta)(8)-barrels that exhibit a previously unseen N-terminal domain swap and have high structural homology …

Total Synthesis Of (-)-Acutumine, Fang Li

Theses and Dissertations

Acutumine is a tetracyclic alkaloid isolated from the Asian vine Menispermum dauricum with selective T-cell cytotoxicity and antiamnestic properties. We have developed a total synthetic route to this congested alkaloid, during which we also found a novel, stereoselective radical-crossover reaction that combines an intramolecular radical conjugate addition with a subsequent enolate hydroxylation. Key features of this synthesis also include a reagent-controlled diastereoselective ketone allylation, an anionic oxy-Cope rearrangement to form a congested quaternary sterocenter, a pyridine-mediated selective ozonolysis, and a Lewis acid promoted Michael-type cyclization.

Development Of Biocompatible Polymer Monoliths For The Analysis Of Proteins And Peptides, Yun Li

Theses and Dissertations

Biocompatibility is an important issue for the development of chromatographic stationary phases for the analysis of biomolecules (including proteins and peptides). A biocompatible stationary phase material is a material that resists nonspecific adsorption of biomolecules and does not interact with them in a way that would alter or destroy their structures or biochemical functions. The monolithic column format is a good alternative to typical spherical particle packed columns for capillary liquid chromatography of biomacromolecules. Several novel anion-exchange polymer monoliths for the analysis of proteins were synthesized for improved biocompatibility. Two novel polymeric monoliths were prepared in a single step by …

Real-Time Virus Analysis Via Image Charge Detection Surface Induced Dissociation Tandem Mass Spectrometry, Seth T. Call

Theses and Dissertations

This thesis reports on the development of a novel mass spectrometer combining image charge detection with surface induced dissociation for real-time analysis of intact viruses. Protonated viruses produced using electrospray are accelerated and subsequently impact on a solid surface. Capsid peptides released during the impact are analyzed using time-of-flight mass spectrometry. Image charge detection is used to measure the mass and charge states of structurally intact, electrosprayed viruses prior to impact. Since virus capsids are composed of loosely-bound proteins, collision of viruses with surfaces at moderate impact energies could release intact proteins. The masses and numbers of different protein types …

Design And Characterization Of A Human Exposure Chamber And Inversion Episodes In Salt Lake City, Utah In January/February Of 2009, Roman Yuri Kuprov

Theses and Dissertations

Research on health effects of particulate matter (PM) has been a very active area in the last two decades. One plausible mechanism by which exposure to PM affects human health includes modification of autonomic endothelium function. Decreased endothelium activity causes heightened risks of cardiovascular disease. A human exposure chamber designed to conduct experiments to quantify diminished function of endothelium from short term exposure to PM is described. The chamber consists of two stages for containment and pre-treatment of PM and exposure of human subjects. Concentrations of CO, CO2, NO, NO2, O3, and PM2.5, are monitored and controlled in the exposure …

The Characterization Of Two Diverse Magnetotactic Bacteria: Lems And Mms-1, Ulysses C. Pickard, Paul A. Howse, Dennis A. Bazylinski

Undergraduate Research Opportunities Program (UROP)

Magnetotactic bacteria (MTB) are a diverse group of prokaryotes that biomineralize membrane-bound magnetic crystals known as magnetosomes. The magnetosomes are aligned within the cell and consist of either magnetite (Fe3O4) or greigite (Fe3S4). The biomineralization of magnetosomes consists of several processes including: invagination of the cytoplasmic membrane, iron uptake into the cell and then into the magnetosome membrane vesicle, and crystallization of the mineral phase inside the vesicle. Mam genes control magnetosome biomineralization with most of the genes present in an island called a magnetosome island. Many of the mam genes are conserved between different species of MTB. The genes …

Geometric Build-Up Solutions For Protein Determination Via Distance Geometry, Robert Tucker Davis

Masters Theses & Specialist Projects

Proteins carry out an almost innumerable amount of biological processes that are absolutely necessary to life and as a result proteins and their structures are very often the objects of study in research. As such, this thesis will begin with a description of protein function and structure, followed by brief discussions of the two major experimental structure determination methods. Another problem that often arises in molecular modeling is referred to as the Molecular Distance Geometry Problem (MDGP). This problem seeks to find coordinates for the atoms of a protein or molecule when given only a set of pair-wise distances between …

The Study Of Variations In The Properties Of Biodiesel On Addition Of Antioxidants, Hiranmayee Kandala

Masters Theses & Specialist Projects

This research studies variations in biodiesel upon addition of different concentrations of antioxidant and petrodiesel additives. Oxidation onset temperature, oxidation induction time, oxidative stability, thermal stability, crystallization onset temperature and moisture retention properties of the biodiesel, with and without antioxidants have been studied. Antioxidants like BHT, BHA, PrG, Vit E and Vit C were added to the biodiesel during the study. These samples were analyzed using the PDSC, TGA, DSC and TGA-SA instrumentation.

The results of this research show an improvement in the OOT, OIT and Oxidative stability of biodiesel with the addition of antioxidants and by blending the biodiesel …

Synthesis And Characterization Of Bis-Phosphine Complexes With Transition Metals, Alicia L. Mcdaniel

Masters Theses & Specialist Projects

Extractants and extraction methodologies play a vital role in many industrial processes, from the concentration of precious metals from ores to the separation of longlived nuclei from radioactive waste as well as the removal of heavy metals from soils and water for remediation. The vast majority of extractants rely on the use of nitrogen, oxygen, sulfur or selenium as Lewis base donor atoms to form coordination complexes with the metal ions of interest. These extractants often make use of the chelate effect and/or the macrocyclic effect in order to form stable complexes. Some of the best known types of chelate …

Cloning And Expression Of A Putative Exported Protein Coding Gene Sas1089 From An Invasive Community Associated Staphylococcus Aureus Strain Mssa476, Nailah Alzaki

All-Inclusive List of Electronic Theses and Dissertations

Originally, methicillin resistant Staphylococcus aureus (MRSA) occurred primarily in the hospital setting and was designated hospital- associated MRSA (HA-MRSA). In recent years, isolates of Staphylococcus aureus from the community (CA-MRSA), found to be methicillin resistant, are not as resistant tooth~ antibiotics as HA-MRSA. However, community strains are noted for their invasiveness causing soft tissue infections with propensity for invasion to deeper sites.

Insulin Stimulates The Phosphorylation Of The Exocyst Protein Sec8 In Adipocytes, Patrick D. Lyons, Grantley R. Peck, Arminja N. Kettenbach, Scott A. Gerber, Liya Roudaia, Gustav E. Lienhard

Dartmouth Scholarship

The signal transduction pathway leading from the insulin receptor to stimulate the fusion of vesicles containing the glucose transporter GLUT4 with the plasma membrane in adipocytes and muscle cells is not completely understood. Current evidence suggests that in addition to the Rab GTPase-activating protein AS160, at least one other substrate of Akt (also called protein kinase B), which is as yet unidentified, is required. Sec8 is a component of the exocyst complex that has been previously implicated in GLUT4 trafficking. In the present study, we report that insulin stimulates the phosphorylation of Sec8 on Ser-32 in 3T3-L1 adipocytes. On the …

An Iterative Strategy Combining Biophysical Criteria And Duration Hidden Markov) Models For Structural Predictions Of Chlamydia Trachomatis S66 Promoters, Ronna R. Mallios, David M. Ojcius, David H. Ardell

All Dugoni School of Dentistry Faculty Articles

Background: Promoter identification is a first step in the quest to explain gene regulation in bacteria. It has been demonstrated that the initiation of bacterial transcription depends upon the stability and topology of DNA in the promoter region as well as the binding affinity between the RNA polymerase σ-factor and promoter. However, promoter prediction algorithms to date have not explicitly used an ensemble of these factors as predictors. In addition, most promoter models have been trained on data from Escherichia coli. Although it has been shown that transcriptional mechanisms are similar among various bacteria, it is quite possible that the …

The Helicobacter Pylori Anti-Sigma Factor Flgm Is Predominantly Cytoplasmic And Cooperates With The Flagellar Basal Body Protein Flha, Melanie Rust, Sophie Borchert, Eike Niehus, Sarah A. Gripp, Afrodita Bajceta, Jonathan L. Mcmurry, Sebastian Suerbaum, Kelly T. Hughes, Christine Josenhans

Faculty Articles

Helicobacter pylori requires flagellar motility and orientation to persist actively in its habitat. A particular feature of flagella in most Helicobacter species including H. pylori is a membraneous flagellar sheath. The anti-sigma factor FlgM of H. pylori is unusual, since it lacks an N-terminal domain present in other FlgM homologs, e.g., FlgM of Salmonella spp., whose regulatory function is intimately coupled to its secretion through the flagellar type III secretion system. The aim of the present study was to characterize the localization and secretion of the short H. pylori FlgM in the presence of a flagellar sheath and to elucidate …

A Top-Down Proteomic Approach For The Discovery Of Novel Serum Biomarkers Of Pregnancy-Related Disease, Karen Merrell

Theses and Dissertations

The serum fraction of blood is an ideal material in which to search for novel biomarkers for disease. It is easily obtained through relatively non-invasive means, routinely collected, and a rich treasure-trove of information about the health of an individual. Cells react to signal molecules, take up nutrients, and release waste products, fragments that are the result of proteolysis, and other molecules out into the bloodstream. If these components are unique to the cells in question, that part of the complex mixture that is the blood stream can potentially characterize the health of the tissue or organ those cells are …

Ion Transport Behaviors Upstream And Downstream From The Sampling Cone Of An Inductively Coupled Plasma Mass Spectrometer, Haibin Ma

Theses and Dissertations

Inductively coupled plasma - mass spectrometry (ICP-MS) is the technique of choice worldwide for trace elemental determinations because of its excellent ionization ability, low detection limits and fast analysis speeds. However, the ICP-MS still suffers from some disadvantages, such as spectral overlap and severe matrix effects. Matrix effects or interferences, partly arise from changes in the analyte transmission through the interfacial region between the ICP and mass spectrometer with changes in sample matrix. Better understanding of the transmission behaviors of analyte through the sampling and skimmer cones will provide the insights needed to alleviate matrix interferences and to improve the …

Relaxation Of Vibrationally Excited Trifluorobenzene And Tetrafluorobenzene By Collisions With Carbon Dioxide, Alan M. Johnson

Theses and Dissertations

An investigation into the relaxation of highly vibrationally excited trifluorobenzene and tetrafluorobenzene following collisions with carbon dioxide was performed using diode laser transient absorption spectroscopy. A 248 nm excimer laser prepared the vibrationally hot (E'~41,000 cm-1) fluorobenzene molecules. Large amounts of translational and rotational energy are transferred through collisions between the hot donor molecule and CO2. Rate constants and collisional probabilities were calculated by probing the high J states (J=58~80) of CO2 in the vibrational ground state, 0000, with measurements taken 1 µsec, ¼ the mean gas collision time, following each excimer laser pulse. The energy transfer probability distribution function, …

The Role Of Smpb In The Early Stages Of Trans-Translation, Deanna June Cazier

Theses and Dissertations

Ribosomes stall on defective messenger RNA transcripts in eubacteria. Without a mechanism to release stalled ribosomes, these cells would die. Transfer-messenger RNA (tmRNA) and small protein B (SmpB) reactivate stalled ribosomes in a process known as trans-translation. Together, tmRNA and SmpB mimic alanyl-tRNA, entering the A site of stalled ribosomes and accepting transfer of the stalled polypeptide. A portion of tmRNA is then positioned as a template for the ribosome to resume translating. The tmRNA open reading frame encodes a proteolysis tag to mark the aberrant polypeptide for degradation and a stop codon to release the ribosome. How are tmRNA …

Micro, Nano Encapsulation Methods For Sustained Release Drug Formulations And Biomimetic Applications, Shantanu Balkundi

Doctoral Dissertations

The Layer-by-Layer (LbL) assembly technique was used to obtain a new type of protein/polyphenol microcapsule based on naturally occurring polyphenol (-)-epigallocatechin gallate (EGCG) and gelatin, type A. The dependence of permeability on the molecular weight of permeating substances was studied and compared with commonly used polyallylamine/polystyrene sulfonate capsules. A quartz crystal microbalance was used to monitor the regularities of EGCG adsorption in alternation with type A and B Gelatins and electrophoretic mobility measurements were used that indicated that the nature of assembly was dependent on Gelatin properties. It was shown that EGCG retains its antioxidant activity in the LbL assemblies. …

Effect Of Chronic Treatment With A Cyclooxygenase Inhibitor On Reproductive Parameters In Male Rat, Sheikh A Saeed, Naheed Anwar, Khalid M Khan, Noorjehan Sarfraz

Department of Biological & Biomedical Sciences

BACKGROUND: Indomethacin is a member of non-steroidal anti-inflammatory drugs (NSAIDs) commonly used for treatment of gout, arthritis, and other inflammatory conditions. It has been shown to inhibit ovarian prostaglandins synthesis in mammals, birds, fish and reptiles. However, the effects of its chronic administration on male reproductive functions remain largely unknown. Using rat as a model, we studied the effect of chronic treatment with indomethacin on the male reproductive system.

METHODS: Testosterone was measured in the serum, testicular tissue, and testicular interstitial fluid by radioimmunoassay. Moreover, we also studied the direct effect of indomethacin in vitro on luteinizing hormone stimulated testosterone …

Inhibition Of Yeast Hexokinase By The Antimalarial Drug Artemisinin: Probing Mechanism Of Action With A Model Enzyme, Jennifer S. Spence

Biological Sciences Theses & Dissertations

A leading infectious cause of death, malaria threatens approximately half of the world's population, and drug-resistant strains of Plasmodium falciparum have created immense difficulty in chemotherapy of the disease. The artemisinin (ART) class of antimalarials may represent a powerful solution. In addition to their safety, effectiveness, and moderate cost, they are the only drugs in use for which there has been no widespread evidence of clinical resistance. The exact parasiticidal mechanism of ART is highly contested, but evidence suggests that protein alkylation may play a role in cytotoxicity. in vitro essays were performed using yeast hexokinase (HK) to demonstrate a …

Functional Characterization Of The Arabidopsis Β-Ketoacyl-Coenzyme A Reductase Candidates Of The Fatty Acid Elongase, Frédéric Beaudoin, Xianzhong Wu, Fengling Li, Richard P. Haslam, Jennifer E. Markham, Huanquan Zheng, Johnathan A. Napier, Ljerka Kunst

Department of Biochemistry: Faculty Publications

In plants, very-long-chain fatty acids (VLCFAs; >18 carbon) are precursors of sphingolipids, triacylglycerols, cuticular waxes, and suberin. VLCFAs are synthesized by a multiprotein membrane-bound fatty acid elongation system that catalyzes four successive enzymatic reactions: condensation, reduction, dehydration, and a second reduction. A bioinformatics survey of the Arabidopsis (Arabidopsis thaliana) genome has revealed two sequences homologous to YBR159w encoding a Saccharomyces cerevisiae β-ketoacyl reductase (KCR), which catalyzes the first reduction during VLCFA elongation. Expression analyses showed that both AtKCR1 and AtKCR2 genes were transcribed in siliques, flowers, inflorescence stems, leaves, as well as developing embryos, but only AtKCR1 …

Applications And Potentials For Biogenic Methane Recovery Operations In Nebraska Agriculture, Industry, And Economic Development, David Micheal Dingman

Department of Environmental Studies: Undergraduate Student Theses

ABSTRACT: This thesis report illustrates the applications and potentials of biogenic methane recovery in Nebraska’s agricultural and industrial sectors and as a means for increasing sustainable economic development in the state’s rural communities. As the nation moves toward a new green economy, biogenic methane recovery as a waste management strategy and renewable energy resource presents significant opportunities for Nebraska to be a national and world leader in agricultural and industrial innovation, advanced research and development of renewable energy technology, and generation of new product markets. Nebraska’s agricultural economy provides a distinct advantage to the state for supporting methane recovery operations …

Novel Cinchona Alkoloid Derived Ammonium Salts As Phase-Transfer Catalysts For The Asymmetric Synthesis Of Beta-Hydroxy Alpha-Amino Acids Via Aldol Reactions And Total Synthesis Of Celogentin C., Bing Ma

Theses and Dissertations

Project I. Cinchona alkaloid-derived quaternary ammonium salts have been successfully used as phase-transfer catalysts, particularly in asymmetric alkylations. Our group applied this type of catalyst in the synthesis of β-hydroxy α-amino acids via aldol reactions and discovered that the Park-Jew catalyst afforded good yields and good enantiomeric excess of the syn diasteromers, but negligible diastereoselectivity. This project was therefore focused on the synthesis of novel cinchonidine-derived catalysts with the Park-Jew catalyst as the lead structure. The C3 position of cinchonidine nucleus was modified to achieve dimers and catalysts possessing electron-deficient alkyne and alkene moieties. Synthesized catalysts were tested in the …