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Structure Of Frequency-Interacting Rna Helicase From Neurospora Crassa Reveals High Flexibility In A Domain Critical For Circadian Rhythm And Rna Surveillance, Yalemi Morales, Keith J. Olsen, Jacqueline M. Bulcher, Sean J. Johnson

Chemistry and Biochemistry Faculty Publications

The FRH (frequency-interacting RNA helicase) protein is the Neurospora crassa homolog of yeast Mtr4, an essential RNA helicase that plays a central role in RNA metabolism as an activator of the nuclear RNA exosome. FRH is also a required component of the circadian clock, mediating protein interactions that result in the rhythmic repression of gene expression. Here we show that FRH unwinds RNA substrates in vitro with a kinetic profile similar to Mtr4, indicating that while FRH has acquired additional functionality, its core helicase function remains intact. In contrast with the earlier FRH structures, a new crystal form of FRH …

Structural Characterization Of The P1+ Intermediate State Of The P-Cluster Of Nitrogenase, Stephen M. Keable, Oleg A. Zadvornyy, Lewis E. Johnson, Bojana Ginovska, Andrew J. Rasmussen, Karamatullah Danyal, Brian J. Eilers, Gregory A. Prussia, Axl X. Levan, Simone Raugei, Lance C. Seefeldt, John W. Peters

Chemistry and Biochemistry Faculty Publications

Nitrogenase is the enzyme that reduces atmospheric dinitrogen (N₂) to ammonia (NH₃) in biological systems. It catalyzes a series of single-electron transfers from the donor iron protein (Fe protein) to the molybdenum–iron protein (MoFe protein) that contains the iron–molybdenum cofactor (FeMo-co) sites where N₂ is reduced to NH₃. The P-cluster in the MoFe protein functions in nitrogenase catalysis as an intermediate electron carrier between the external electron donor, the Fe protein, and the FeMo-co sites of the MoFe protein. Previous work has revealed that the P-cluster undergoes redox-dependent structural changes and that the …

Small-Angle X-Ray Scattering In Amorphous Silicon: A Computational Study, Durga Paudel, Raymond Atta-Fynn, David A. Drabold, Stephen R. Elliott, Parthapratim Biswas

Faculty Publications

We present a computational study of small-angle x-ray scattering (SAXS) in amorphous silicon (α-Si) with particular emphasis on the morphology and microstructure of voids. The relationship between the scattering intensity in SAXS and the three-dimensional structure of nanoscale inhomogeneities or voids is addressed by generating large high-quality (α-Si networks with 0.1%–0.3% volume concentration of voids, as observed in experiments using SAXS and positron annihilation spectroscopy. A systematic study of the variation of the scattering intensity in the small-angle scattering region with the size, shape, number density, and the spatial distribution of the voids in the networks is presented. Our results …

Functional And Biochemical Characterization Of Dib1'S Role In Pre-Messenger Rna Splicing, Christian C. Schreib, Emily K. Bowman, Cody A. Hernandez, Amber L. Lucas, Camile H. S. Potts, Corina Maeder

Chemistry Faculty Research

The spliceosome is a dynamic macromolecular machine that undergoes a series of conformational rearrangements as it transitions between the several states required for accurate splicing. The transition from the B to Bact is a key part of spliceosome assembly and is defined by the departure of several proteins, including essential U5 component Dib1. Recent structural studies suggest that Dib1 has a role in preventing premature spliceosome activation, as it is positioned adjacent to the U6 snRNA ACAGAGA and the U5 loop I, but its mechanism is unknown. Our data indicate that Dib1 is a robust protein that tolerates incorporation of …

Molecular Dynamics Simulations Of Apo And Holo Forms Of Fatty Acid Binding Protein 5 And Cellular Retinoic Acid Binding Protein Ii Reveal Highly Mobile Protein, Retinoic Acid Ligand, And Water Molecules, Nathanael H. Hunter, Blair C. Bakula, Chrystal D. Bruce

2018 Faculty Bibliography

Structural and dynamic properties from a series of 300 ns molecular dynamics, MD, simulations of two intracellular lipid binding proteins, iLBPs, (Fatty Acid Binding Protein 5, FABP5, and Cellular Retinoic Acid Binding Protein II, CRABP-II) in both the apo form and when bound with retinoic acid reveal a high degree of protein and ligand flexibility. The ratio of FABP5 to CRABP-II in a cell may determine whether it undergoes natural apoptosis or unrestricted cell growth in the presence of retinoic acid. As a result, FABP5 is a promising target for cancer therapy. The MD simulations presented here reveal distinct differences …

Discovery Of A Novel Inhibitory Compound Produced By The Soil Bacterium Rhodococcus Sp. Mtm3w5.2, Patrick South

Undergraduate Honors Theses

Due to increases in antibiotic resistance stemming from the overuse of commercial antibiotics, the need to discover novel antibacterial compounds is becoming more urgent. A species of Rhodococcus, MTM3W5.2, has been discovered and was found to produce a metabolite with inhibitory activity against closely related species. The aim of this study is to elucidate the structure of the inhibitory metabolite by isolating and purifying it; then characterizing it using spectroscopic techniques. The compound was isolated from MTM3W5.2 RM broth cultures using n-butanol extraction, which yielded an active crude extract. The crude extract was then subjected to fractionation using a Sephedex …

Approaches Toward The Total Synthesis Of Oroidin Alkaloids:Nagelamide A, C And Ageliferin, Apsara Herath

Chemistry & Biochemistry Dissertations - Archive

Nagelamides and ageliferin are oroidin-derived alkaloids which have gained the attention of the scientific community, because of their significant biological activity as well as the unprecedented structural diversity. Our work is directed toward the development of synthetic methods for the construction of oroidin dimers. Accordingly, this dissertation is composed of two sections; the chapter one provides an overview of the oroidin alkaloids with a particular focus on the nagelamide family and the third chapter describes synthetic studies towards ageliferin. The first part of chapter two focuses on the Palladium-catalyzed Stille cross-coupling reaction for the construction of the main dimeric scaffolds …

Decreased Brood-Size Of Starvation-Induced Postdauer Adults Is Inherited Transgenerationally Via Hrde-1, Ibnul Rafi

Renée Crown University Honors Thesis Projects - All

Environmental stress experienced in utero or in early life has profound effects on the behavior and physiology of an adult animal. Interestingly, genome-wide association studies in humans have shown that early-life stress not only affects the exposed individual, but also their future generations. However, the molecular mechanisms of how early-life stress can impact physiology transgenerationally in the absence of the original stress are not well understood. To study the mechanisms and consequences of environmental stress on adult phenotypes, we utilize the life cycle of the model organism Caenorhabditis elegans. In favorable conditions, the life cycle of C. elegans includes 5 …

Supercharging And Multiple Reaction Monitoring Optimization Of High Molecular Weight Intact Proteins Using Triple Quadrupole Mass Spectrometry, Durga Devi Khanal

Chemistry & Biochemistry Theses - Archive

Various supercharging agents, such as meta-nitrobenzyl alcohol, dimethyl sulfoxide, trifluoroethanol (TFE), and sulfolane were tested in different concentrations to study the effects of supercharging agents on the charge distributions of the high molecular weight intact proteins. The data were analyzed by triple quadrupole mass spectrometry. For most of the proteins, m-NBA and TFE worked more effectively than other supercharging agents, both to shift the charge state and increase intensity. The settings of different instrumental parameters were also studied. Both increases in source temperature and flow rate had a negative impact on average charge state observed. The goal of this work …

Spectroscopic And Mechanistic Investigation Of The Enzyme Mercaptopropionic Acid Dioxygenase (Mdo), The Role Of The Conserved Outer-Sphere 'Catalytic Triad' In Directing Promiscuous Substrate-Specificity, Sinjinee Sardar

Chemistry & Biochemistry Dissertations - Archive

Mercaptopropionate dioxygenase (MDO) from Aztobacter Vinelandii is a non-heme mononuclear iron enzyme that catalyzes the oxygen dependent conversion of 3-mercaptopropionic acid (3mpa) to produce 3-sulfinopropinioic acid (3spa). Nearly, all thiol dioxygenases have a conserved ‘catalytic triad’ within the active site, consisting of three outer Fe-coordination sphere residues S155, H157 and Y159. With the notable exception of mammalian cysteamine (2-aminoethanethiol) dioxygenase (ADO), this ‘catalytic triad’ is universally conserved across phylogenic domains. X-ray crystallographic studies clearly show a hydrogen bond network connecting the ‘catalytic triad’ to the mononuclear iron site; however, its influence on substrate specificity and catalytic efficiency is poorly understood. …

Elucidation Of Substrate Binding To Photolyase From Sulfolobus Solfataricus Using Isothermal Titration Calorimetry, Amanda Rose Kuerzi

Theses, Dissertations and Culminating Projects

DNA photolyase is a DNA repair enzyme commonly found across the kingdom of life. Binding studies for the hyperthermophilic photolyase derived from Sulfolobus solfataricus (SsPL) may serve to illuminate how DNA photolyase can adapt over a large temperature range. In the first section of this study, the thermodynamics of substrate binding for SsPL in a choline chloride buffer were determined using isothermal titration calorimetry. Using the Counter-Ion Condensation Concept as a model, ionic strength studies were performed to separate the binding interactions into electrostatic, and nonelectrostatic components. The electrostatic interactions do not appear to make a significant contribution to the …

Forensic Identification Of Condom Lubricants By Advanced Statistical Analysis Processing Of Direct Analysis In Real Time Mass Spectrometric Data – Towards Development Of A Condom Residue Database For Forensic Science Practitioners, Allix M. Coon

Chemistry

The ability to identify condom derived trace evidence is gaining importance because of the increasing frequency with which perpetrators of sexual assault use condoms in order to avoid leaving behind incriminating DNA evidence that might link them to the crime. Although DNA remains the gold standard for sexual assault evidence, other forms of trace evidence are needed in its absence. When condoms are used, the lubricants associated with them are the trace evidence. For the lubricant residue to be useful in a forensics context, a database of chemical signatures of lubricants against which acquired evidence can be screened is required, …

Evaluation Of The Biocompatibility And Micromechanical Properties Of Experimental Dental Adhesives, Hoda Moussa

Forensic Science Theses

The purpose of this study was to investigate the biocompatibility and micromechanical properties of newly synthesized antibacterial monomer and cross-linker functional surfactants into a commercially available dental adhesive (Single Bond, 3M ESPE, Saint Paul, MN, USA) at three concentrations (0.05, 0.1, 0.25 mg/mg). All groups were analyzed by evaluation of micro-tensile bond strength, ultimate tensile strength, cell viability, antibacterial properties, and surface micro-hardness. Scanning electron microscopy (SEM) was used for interfacial characterization.

Human extracted molars were used as a substrate for bonding adhesives for the micro-tensile bond strength (MTBS) and scanning electron microscopy (SEM) studies. Twenty resin-dentin beams (0.9 ± …

An Exploration Of Pyrrole Groups On The Enantioselectivity Of Pig Liver Esterase, Brian Long

Honors Theses

Pig Liver Esterase (PLE) is a serine protease enzyme that can interact with one side of a diester to hydrolyze the ester to a carboxylic acid, and research has found that the level of hydrophobicity of side groups can impact the enantioselectivity of PLE hydrolysis.1, 2 The Jones Model is what current researchers use to model the active site of PLE, but the nature of its binding pockets, namely the Hydrophobic Long (HL) pocket, has been called into question.3 Dimethyl 2-((pyrrole-2-yl)methyl)-2-methylmalonate was prepared to be subjected to PLE hydrolysis to see whether enantioselectivity was found. Chiral HPLC revealed 25.32% enantiomeric …

Pro-Antimicrobial Networks Prepared Via Degradable Thiol-Ene Acetal Photopolymerization, Sarah Swilley

Honors Theses

A degradable, antimicrobial polymer network with acetal crosslink junctions derived from p-anisaldehyde (pA), a common constituent found in star anise extract, is reported. The p-anisaldehyde was converted into a bis-functional acetal alkene monomer, which serves as a pro-antimicrobial form of the active aldehyde and a building block for materials referred to as pro-antimicrobial polymer networks via degradable acetals (PANDAs). Subsequently, the bis-functional acetal alkene monomer was photopolymerized with a multifunctional thiol to yield a thiol-ene network. PANDAs exhibit surface erosion behavior and yield sustained release of pA over 38 days when exposed to neutral or biologically relevant conditions. The pA …

Diastereoselective Synthesis Of 2,4,6-Trisubstituted Piperidines Via Aza-Prins Cyclization, John A. Hood

Honors Theses

The nitrogen heterocycles are shared amongst 59% of Food and Drug Administration (FDA) approved small molecule pharmaceuticals with the six-membered piperidine representing the most common moiety. Given the versatility and potential to yield derivatives with broad biological activities, the discovery of new chemical methods to generate these heterocycles in a more time and cost-efficient manner is desired. While there are existing racemic methods to access this class of molecule, the objective of this research is to pioneer a new novel six-step method to generate 2,4,6-trisubstituted piperidines with stereoselective control.

The first step is a condensation between a nonenolizable aldehyde and …

Development Of Microdialysis Probes In Series Approach Toward Eliminating Microdialysis Sampling Calibration: Miniaturization Into A Pdms Microfluidic Device, Randy Espinal Cabrera

Graduate Theses and Dissertations

A new microdialysis sampling method and microfluidic device were developed in vitro. The method consisted of using up to four microdialysis sampling probes connected in series to evaluate the relative recovery (RR) of different model solutes methyl orange, fluorescein isothiocyanate (FITC)-dextran average mol. wt. 4,000 (FITC-4), FITC-10, FITC-20, and FITC-40. Different flow rates (0.8, 1.0, and 1.5 µL/min) were used to compare experimentally observed relative recoveries with theoretical estimations. With increasing the number of probes in series, the relative recovery increases and ~100% (99.7% ± 0.9%) relative recovery for methyl orange was obtained. For larger molecules such as fluorescein isothiocyanate …

Computational Modeling Of Electronically Excited States In Cobalamin-Dependent Reactions., Brady D. Garabato

Electronic Theses and Dissertations

The current understanding of the photolytic properties of Vitamin B12 derivatives or cobalamins are summarized from a computational point of view. The focus is on two non-alkylcobalamins, cyanocobalamin (CNCbl) and hydroxocobalamin (HOCbl), two alkylcobalamins, methylcobalamin (MeCbl) and adenosylcobalamin (AdoCbl), as well as the stable cob(II)alamin radical. Photolysis of alkylcobalamins involves low-lying singlet excited states where photo-dissociation of the Co-C bond forms singlet-born alkyl/cob(II)alamin radical pairs (RPs). Potential energy surfaces (PESs) of low-lying excited states as functions of both axial bonds provide the most reliable tool for analysis of photochemical and photophysical properties. Due to the size limitations associated with the …

The Effects Of Passive Integrated Transponder Tagging On Cortisol Release In The Gulf Killifish, Fundulus Grandis, Ariel Marcelo Hernandez

Senior Honors Theses

Due to its ease of use, low cost, and essentially limitless number of unique codes, PIT tagging has become the favored technique for tagging individuals in biological studies. However, studies employing PIT tagging generally assume that stress due to the implantation and presence of a PIT tag has no influence on the experimental results. This study investigated the effects of PIT tagging on levels of the stress hormone cortisol in the Gulf Killifish, Fundulus grandis, an estuarine fish of the Gulf of Mexico that is prone to daily or seasonal environmental stressors. Cortisol was measured non-invasively by extracting and …

Ultrafast Affinity Extraction And High-Performance Affinity Chromatography Applications For Measuring Free Drug Fractions: Interactions Of Sulfonylurea Drugs With Normal And Glycated Human Serum Albumin, Bao Yang

Department of Chemistry: Dissertations, Theses, and Student Research

Ultrafast affinity extraction was used with microcolumns containing human serum albumin (HSA) to measure the free fractions of several sulfonylurea drugs in the presence of solution-phase HSA and for the analysis of these drugs at therapeutic levels or determining their binding strengths with this protein. This approach was first used with normal HSA and the drugs glibenclamide, glipizide and glimepiride. This work used both one- and two-dimensional affinity systems based on HSA microcolumns. The first microcolumn was used to extract the free drug fractions in injected drug/protein mixtures. As the retained drug eluted from the first microcolumn, it was passed …

Phospholipid Membrane Remodeling Through Exogenous Fatty Acid Incorporation In Certain Bacterial Species, Josh Herndon

Honors Theses

Multi-drug resistant bacteria have become problematic in healthcare settings. Recent studies have documented the ability of some bacteria, such as Vibrio species, to assimilate exogenous fatty acids into their membrane phospholipids. Therefore, the current research was performed to determine whether the three virulent, multi-drug resistant bacteria Acinetobacter baumannii, Klebsiella pneumoniae, and Escherichia coli can remodel their cell membranes following exposure to a range of exogenous polyunsaturated fatty acids (PUFAs). Furthermore, we hypothesized that PUFAs would influence phenotypes associated with growth and virulence. Extracted phospholipids were examined by UPLC/MS analysis to confirm assimilation of exogenous PUFAs into the bacterial phospholipids. A. …

Sds-Page Analysis Of Modified Lysozyme By Substituted Benzoquinones And Naphthoquinones, Neethu Kurien

Honors Theses

Quinones are molecules that are commonly found in the environment, as cofactors for proteins or an electron carrier in the mitochondrial electron transport chain. However, quinones also can come from polycyclic aromatic hydrocarbons as their metabolites, harming cellular integrity. In this study, we investigated protein modification of lysozyme in the presence of substituted benzoquinones and naphthoquinones. To simulate physiological conditions, all reactions used phosphate buffer (50.0 mM). While the majority of the experiments were performed at 37 C and pH 7.0, there were also temperature dependence (27 C and 42 C) and pH dependence (6.0 and 8.0) reactions included. SDS-PAGE …

Development Of New Useful Methods For Aldehyde Synthesis And Its Application In Synthesis, He Huang

Chemistry and Chemical Biology ETDs

Development of sustainable synthetic technologies for molecular construction is an important but formidably challenging task in modern organic synthesis. Aldehyde synthesis represents a long-standing interest in synthesis because of the synthetic utility. Classic methods for aldehyde synthesis have the drawbacks of the use of harsh reaction conditions, poor atom-economy and multi-step operation, and production of stoichiometric amount of chemical wastes. The state-of-the-art strategies employ transition metal complexes as catalysts to promote formylation reactions. The concerns of catalyst cost, operation complexity and poor functional group tolerance demands more efficient synthetic technologies. My Ph.D. study focuses on design of conceptually novel catalytic …

Singlet Oxygen Oxidation Of Guanine, 9-Methylguanine And Guanine-Cytosine Base Pair: Dynamics And Kinetics Revealed By Parallel Gas- And Solution-Phase Experiments And Computations, Wenchao Lu

Dissertations, Theses, and Capstone Projects

Singlet oxygen (¹O₂) oxidatively generated damage of DNA gives rise to mutagenesis, carcinogenesis, and cellular death. Guanine is the most susceptible DNA target of ¹O₂. The related process has been studied over three decades but the mechanism has remained elusive. My thesis research has focused on reaction mechanism, dynamics and kinetics of ¹O₂ oxidation of guanine, 9-methylguanine and guanine-cytosine base pair, from the gas-phase bare ions, through hydrated clusters, to aqueous solution. Various techniques have been adapted in the work, including ¹O₂ generation and detection, guided-ion beam tandem mass …

Removal Of Chromium(Vi) And Chromium(Iii) Ions From Aqueous Solution Using Bio-Char Generated From Agricultural Waste Products, Diego Fernando Gonzalez

Theses and Dissertations

Heavy metals are one of the most persistent and prevalent contaminants in the aquatic environment. The removal of chromium from aqueous solution, especially in the hexavalent form is difficult. New technologies, techniques and/or new materials have been designed in order to effectively and efficiently remove chromium from the aqueous environment. The project focuses on the comparison of the effects of pH, time, temperature, binding capacity on bio-char’ generated from agricultural waste produces vs an amino modified derivative of the bio-char. Pineapple skins were dried, ground, sieved, and pyrolyzed to produce a bio-char material. The bio-char was analyzed using FTIR to …

Elucidating Molecular Interactions Of Shigella Type Three Secretion System Components Critical For Pathogenesis, R. Alan Burgess

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Shigella are pathogenic bacteria which commonly infect human hosts through the fecal-oral route. As the causative agent of shigellosis, a disease characterized by severe diarrhea, nausea, and vomiting, Shigella are responsible for an estimated 90 million infections and 100,000 deaths each year. With the emergence of antibiotic resistant strains of Shigella, the need for alternative treatments methods has become critical to fighting outbreaks of shigellosis and drug resistant strains. One such method is to target the infection system of the bacteria, called the type three secretion system (T3SS). The T3SS includes a complex needle and syringe-like apparatus that is …

Investigations Into Ring-Opening Polymerization Of Functionalized Ε-Caprolactones, Danielle N. Coderre, Matthew K. Kiesewetter

Senior Honors Projects

Polymers are defined as large molecules constructed from small, repeating units (monomers) which make up everyday items, such as: plastic bags and bottles, waterproof coatings, pipes, etc. H-bonding urea (U) or thiourea (TU) catalysts paired with base cocatalysts used for organocatalytic ring-opening polymerization (ROP) of cyclic esters have been shown to exhibit fast rates of polymerization and result in polymers with controlled molecular weights (Mn) and low polydispersity indexes (PDI). Additionally, organocatalytic ring-opening polymerization (ROP) techniques have been shown to be tolerant of the incorporation of functional groups into the monomer feed, which is vital for the catalytic synthesis of …

Multivalency Regulates Activity In An Intrinsically Disordered Transcription Factor, Sarah Clark, Janette B. Myers, Ashleigh King, Radovan Fiala, Jiri Novacek, Grant Pearce, Steve L. Reichow, Elisar J. Barbar

Chemistry Faculty Publications and Presentations

The transcription factor ASCIZ (ATMIN, ZNF822) has an unusually high number of recognition motifs for the product of its main target gene, the hub protein LC8 (DYNLL1). Using a combination of biophysical methods, structural analysis by NMR and electron microscopy, and cellular transcription assays, we developed a model that proposes a concerted role of intrinsic disorder and multiple LC8 binding events in regulating LC8 transcription. We demonstrate that the long intrinsically disordered C-terminal domain of ASCIZ binds LC8 to form a dynamic ensemble of complexes with a gradient of transcriptional activity that is inversely proportional to LC8 occupancy. The preference …

Polymer Nanocomposites Containing High Aspect Ratio Particulates: Innovation In Co-Extruded Multilayer Barrier Films, Kevin Meyers

Dissertations

Delaminated montmorillonite (MMT) clay/ maleic anhydride grafted LLDPE nanocomposite multilayer films with alternating layers of LDPE were produced through multilayer co-extrusion. The MMT concentration within the nanocomposite layers was increased through annealing the films in the melt due to a mismatch in interdiffusion rates of the polymer layers. Analysis of the nanocomposite layers upon annealing revealed that the platelets impinged upon one another resulting in significant improvement in oxygen barrier in the multilayer system, exceeding the results of bulk nanocomposites.

Model analysis demonstrated that increasing the nanoplatelet aspect ratio or initial concentration in the filled layers would lead to even …

Evaluation Of Cafo Dairy Manure Treatment To Reduce Nutrient Transport, Jenna Laib

Honors Projects

Excess amounts of phosphorus and nitrogen flowing into Lake Erie from agricultural fields in Northwest Ohio has led to several harmful algal blooms (HABs). One potential source of those nutrients is manure applied to fields for fertilizer. Manure from confined animal feeding operations (CAFOs) is 95-98% water with only ~3% solids and nutrients, thus physical transportation is expensive relative to the value of the agricultural nutrients. Furthermore, once manure nutrients are applied to agricultural fields, they are relatively easily mobilized to waterways by precipitation. More than 800 lab-scale tests have been used to optimize the treatment of CAFO manure with …