Biochemistry Commons^™

Structural Features Of Hiv-1 Integrase In The Late Stage Of Viral Replication, Tolga Catmakas

Dissertations

HIV-1 integrase (IN) is an essential enzyme for the viral life cycle as it integrates viral genomic material into the host cell genome. In addition to its’ essential catalytic function, IN has another important non-catalytic role where it binds the viral RNA to facilitate successful particle maturation during the viral life cycle. A significant number of IN mutants, addressed as class II IN mutants, show various defects, affecting different steps of the viral life cycle. Class II IN mutants exhibit aberrant morphology where viral ribonucleoprotein particles are mislocalized outside of the capsid core. This phenomenon can be observed in the …

Hiv-1 Vpu Protein Forms Stable Oligomers In Aqueous Solution Via Its Transmembrane Domain Self-Association, Saman Majeed, Lan Dang, Md Majharul Islam, Olamide Ishola, Peter P Borbat, Steven J Ludtke, Elka R Georgieva

Faculty, Staff and Students Publications

We report our findings on the assembly of the HIV-1 protein Vpu into soluble oligomers. Vpu is a key HIV-1 protein. It has been considered exclusively a single-pass membrane protein. Previous observations show that this protein forms stable oligomers in aqueous solution, but details about these oligomers still remain obscure. This is an interesting and rather unique observation, as the number of proteins transitioning between soluble and membrane embedded states is limited. In this study we made use of protein engineering, size exclusion chromatography, cryoEM and electron paramagnetic resonance (EPR) spectroscopy to better elucidate the nature of the soluble oligomers. …

Mechanisms Of Fibrillar Aggregation Of The Lens Small Heat Shock Proteins, Αa- And Αb-Crystallin, Russell James Mcfarland

Dissertations and Theses

The protein αB-crystallin represents the archetypical small heat-shock protein (sHSP), and together with αA-crystallin makes up the primary chaperone system in the eye lens, where they play a critical role in maintaining proteostasis. To achieve transparency, the lens undergoes a loss of organelles and both protein turnover halts, requiring that proteins remain functional for long periods of time as they accumulate chemical damage. The α-crystallins are highly abundant in the lens to stabilize proteostasis for multiple decades. Over time though, the small heat-shock proteins become overwhelmed by a high burden of destabilized clients and their own accumulated chemical damage. Mechanistic …

Institutional And Individual Approaches To Change In Undergraduate Stem Education: Two Framework Analyses, Stephanie B. Feola

USF Tampa Graduate Theses and Dissertations

The overarching goal of this dissertation is to examine institutional change initiatives in STEM and to understand the impact of engaging with a specific change strategy on a biochemistry educator’s instructional practice. Institutional change initiatives involve executing an intentional strategy to influence teaching practices within a specific institution. This research investigates NSF-funded change initiatives at three public research universities that were focused on encouraging the adoption of evidenced-based instructional practices (EBIPs) by STEM faculty. As part of an effort to understand how the three institutions approached this task, framework analysis was performed using the initiative’s annual reports to NSF. Further …

First Course Portfolio For Bioc205: Scientific Analysis And Technical Writing, Lindsey B. Crawford

UNL Faculty Course Portfolios

BIOC205: Scientific Analysis and Technical Writing is a core course for the Department of Biochemistry at the University of Nebraska-Lincoln. This course is geared for second year (sophomore students) with minimal prerequisites and as a foundation for future classes. Challenges include 1) teaching the needed scientific background and elements of scientific analysis and writing to a broad cohort of students with diverse backgrounds and interests, 2) building student skills towards analysis of primary peer-reviewed scientific literature, 3) developing students as writers of scientific information for different audiences. This course used Genetic Engineering as a broad topic choice to capture students …

A Synthetic Approach To The Optimization Of Silicon-Rosindolizine Swir Fluorophores, Timothy Lewis

Honors Theses

Fluorophores emitting within the near-infrared (700 – 1000 nm) or shortwave-infrared (1000 – 1700 nm) regions of the electromagnetic spectrum are of particular interest for their potential to supplement or even outright replace commonly used biological imaging techniques, such as magnetic resonance imaging and computed tomography scanning. Various characteristics make these molecules particularly appealing to researchers, such as their ease of synthetic tunability, generally low toxicity, and high solubility in biological media in comparison to other NIR- and SWIR-emissive fluorophores. Different molecular skeletons have been examined for use as the core of these imaging agents, such as squaraine, cyanine, and …

Extension Of The Ergot Alkaloid Gene Cluster, Samantha Joy Fabian

Graduate Theses, Dissertations, and Problem Reports (ETD)

Specialized metabolites produced by fungi impact human health. A large portion of the pharmaceuticals currently on the market are derived from metabolites biosynthesized by microbes. Ergot alkaloids are a class of fungal metabolites that are important in the interactions of environmental fungi with insects and mammals and also are used in the production of pharmaceuticals. In animals, ergot alkaloids can act as partial agonists or antagonists at receptors for 5-hydroxytryptamine (serotonin), dopamine, and noradrenaline as ergot alkaloids have chemical structures similar to those neurotransmitters. Therefore, they affect insects and mammals that consume them and can be used to produce drugs …

Development Of Multivalent Dna-Peptide Nucleosome Mimetics And Multi-Domain Protein Inhibitors That Directly Or Indirectly Target The E3 Ligase Uhrf1, Li Gu

University of the Pacific Theses and Dissertations

UHRF1 is an E3 ubiquitin ligase and a key epigenetic regulator establishing a crosstalk between DNA methylation and histone modification. Despite the important biochemical role of UHRF1 in cells, its overexpression has been found in almost all primary cancer types including breast cancer, lung cancer and so on. Numerous evidence indicates a strong link between tumorigenesis and UHRF1 overexpression, supporting its potential as a universal biomarker for cancer. However, UHRF1 is “yet-to-be drugged” and no highly potent chemical probes have been developed to target UHRF1 to date. In this study, we proposed two drug design approaches for UHRF1. The first …

Functional Residues Of Cjpata, Robert Lamont

Theses and Dissertations (Comprehensive)

Antibiotic resistance in pathogenic bacteria continues to challenge clinicians and threaten the lives of infected individuals. For this reason, new classes of antibiotics or new targets for antibiotic therapy are needed to circumvent this global health crisis. In Gram-negative bacteria, the enzymes peptidoglycan O-acetyltransferase A and B (PatA and PatB), are responsible for adding acetyl groups to the C-6 hydroxyl group of N-acetyl muramic acid (MurNAc) in peptidoglycan (PG). These acetyl groups are responsible for inhibiting the activity of lysozyme from host organisms that hydrolyze the β1-4 glycosidic linkage between MurNAc and N-acetylglucosamine (GlcNAc). The acetyl groups …

Development Of Nucleic Acid Diagnostics For Targeted And Non-Targeted Biosensing, Christopher William Smith

Legacy Theses & Dissertations (2009 - 2024)

The field of nucleic acid technology is rapidly expanding with new impactful discoveriesbeing made each year. Starting from the discovery of the double-helix structure, cloning, gene editing, polymerase chain reaction (PCR), CRISPR technology, and even the late mRNA vaccines; nucleic acid technology is at the forefront of improving medicine. Nucleic acid technology is extremely versatile due to its easy programmability, automated cheap synthesis, and even its catalog for numerous chemical modifications that can be used to alter structure stability. For example, the number of permutations that can be made with DNA just by altering the code for adenine (A), cytosine …

Molecular Characterization Of Nitrogenase Regulation In Methanosarcina Acetivorans, Melissa Chanderban

Graduate Theses and Dissertations

Nitrogenase is the metalloenzyme only found in bacteria and archaea that is essential for biological nitrogen fixation (diazotrophy), but it can also serve as a catalyst in biofuel production. All diazotrophs contain a molybdenum (Mo) nitrogenase, while some species contain additional alternative nitrogenases where either vanadium (V) or iron (Fe) replace Mo in the active site cofactor. Nitrogen fixation by bacteria has been extensively studied. The limited investigation of nitrogen fixation in methanogenic archaea (methanogens) indicates production of nitrogenase is simpler than in bacteria and methanogen nitrogenase has different biochemical properties. Thus, methanogen nitrogenases provide a promising alternative for genetic …

Expression And Purification Of E. Coli Yoaa, A Putative Helicase, Mark Gregory, Vincent Sutera Mr., Susan Lovett Dr.

Medical Student Research Symposium

All cells must maintain their genomic integrity to survive, which they achieve through several repair mechanisms that necessitate unwinding the damaged DNA by helicases. In Escherichia coli (E. coli), YoaA has been genetically shown to be involved in DNA repair and shares conserved sequences with helicase DinG. The goal of our study was to purify YoaA for further biochemical characterization. For expression, YoaA was fused to a His tag and overexpressed in MG1655 E.coli under the lacZ or T7 promoters for 2 hours, 4 hours, or overnight at 24oC, 30oC or 37oC. For purification, crude lysate was applied to a …

Targeting Heat Shock 27 Kda Protein Induces Androgen Receptor Degradation, Yaxin Li

ETD Archive

Glioblastoma (GBM) is the most common and aggressive brain tumor, with very poor prognosis. Androgen receptor (AR) plays a significant role in the progression of GBM, and anti-androgen agents have the potential to be used for the treatment of GBM. However, AR mutation commonly happens in GBM, which makes the anti-androgen agents less effective. Heat shock 27 kDa protein (HSP27) is a well-documented chaperone protein to stabilize AR. Inhibition of HSP27 results in AR degradation regardless the mutation status of AR, which makes HSP27 a good target to abolish AR in GBM. Identified compound I ((N-(3-((2,5-dimethoxybenzyl)oxy)-4-(methylsulfonamido) phenyl)-4-methoxybenzamide) inhibits GBM cell …

Assembly Of The Peripheral Arm Subunits Of Escherichia Coli Complex I And Analysis Of Clinical Mutations, Hind Alkhaldi

Biological Sciences Theses and Dissertations

Respiratory Complex I from E. coli is a proto-type of the mitochondrial enzyme, consisting of a 6-subunit peripheral arm (B-CD-E-F-G-I) and a 7-subunit membrane arm. When subunits E-F-G (N-module), were expressed alone they formed an active complex as determined by co-immunoprecipitation and native gel electrophoresis. When co-expressed with subunits B and CD, only a complex of E-F-G was found. When these five subunits were co-expressed with subunit I and two membrane subunits, A and H, a complex of B-CD-E-F-G-I was membrane-bound, constituting the N- and Q-modules. Assembly of Complex I was also followed by splitting the genes between two plasmids, …

Computational Investigation Of Calmodulin Photocontrol With The Help Of An Azobenzene Derivative, Jeremy Wells

Undergraduate Honors Theses

The ability to control the activity and binding capability of enzymes in a reversible manner offers tremendous control over biological processes. Photocontrol, in particular, is promising in that electromagnetic radiation can be fine-tuned in terms of its strength, location, and duration. Photosensitive compounds, such as the azobenzene family, experience an isomerization at certain wavelengths, and attaching these compounds to enzymes has the potential to alter their structure and activity. Calmodulin (CaM) is a Ca2+ -sensitive signaling protein that has the ability to affect several downstream processes in eukaryotes and is an excellent target for photocontrol due to its small size …

Mechanisms Of Telomere Maintenance In Trypanosoma Brucei, M A G G. Rabbani

ETD Archive

Telomeres are a nucleoprotein structure at the end of the chromosome and are essential for genome integrity and chromosome stability. Telomere lengths are primarily maintained by a telomerase-mediated pathway but can be maintained by a homologous recombination-mediated pathway. However, detailed mechanisms of telomere maintenance are still unclear in many eukaryotes, including an important human pathogen, Trypanosoma brucei. Telomeres can be elongated by telomerase in T. brucei, a causative agent of fatal sleeping sickness in humans and nagana in cattle. T. brucei evades host immune response by regularly switching its major surface antigen, variant surface glycoprotein (VSG), a process known as …

Covalent Modification Of Recombinant Protein With Reactive Thiols, Sawyer Dulaney, Bailey Taylor

Honors Theses

Many diseases cause chronic and painful inflammation in different body systems. One of the front-line drug classes to treat such inflammation is Nonsteroidal Anti-Inflammatory Drugs (NSAIDs). Despite the benefits of oral administration of NSAIDs, there are drawbacks to their long-term usage because they can cause detrimental effects on off-target systems in the body such as the liver, kidney, or the lining of the intestinal tract. An alternative to NSAIDs is the usage of hydrogels for targeted drug delivery. Hydrogels can provide drug delivery in a specific portion of the site of inflammation, thus allowing higher doses of medication to be …

Targeting Platelets Using Ionic Liquid Coated Nanoparticles, Karen Mun Wong

Honors Theses

Platelets are key players in our body’s inflammatory processes upon activation, which can create physiological problems. Researchers are actively seeking solutions by targeting platelets with therapeutics. The objective of the research in this thesis was to investigate a new and potentially less invasive drug delivery platform using nanoparticles. The topic of the project was to explore the world of ionic liquids and their abilities, when coated on nanoparticles, to bind to platelets. A library of choline carboxylic acid-based ionic liquids (ILs) was first synthesized via salt metathesis while PLGA-based nanoparticles were simultaneously being synthesized via the nanoprecipitation method. The ionic …

Investigation Of The Role Of Overexpression Of Psbs Under Stress Inducible And Constitutive Promoters To Improve Water Use Efficiency., Annie Nelson

Honors Program: Senior Projects (Public)

As climate change continues to impact environmental growth conditions, it has become increasingly more important to identify potential mechanisms of crop development to resist these changes. Previous studies have identified the role of PsbS in the non-photochemical quenching (NPQ) mechanism in the plant by identifying its direct effect on the rate in which excitation energy absorbed by photosystem II is dissipated as heat. It was identified also that PsbS via NPQ oxidizes chloroplastic quinone A (QA) which is a signal for stomatal opening in response to light. By identifying this relationship between PsbS and the signal for stomatal opening in …

Investigating Spatiotemporal Kinetics, Dynamics, And Mechanism Of Exosome Release, Anarkali Mahmood

Electronic Theses and Dissertations

Exosomes are small lipid-based vesicles that can carry biomolecules from one cell to another. While exosomes are crucial to maintain homeostasis in healthy cells, they are exploited by unhealthy cells to aid disease progression. Exosomes likely facilitate disease progression via the transfer of disease-causing biomolecules from unhealthy to healthy cells. Exosomes are generated in Multivesicular endosomes (MVEs) and are then secreted into the extracellular space to travel to other cells. Despite being a crucial step, very little is known about exosomes release mechanism and dynamics. To further our understanding of exosomes, specifically their secretion, my work has focused on investigating …

Investigation Of A Carbon Monoxide Dehydrogenase From An Uncultured Archaeon, Luke Moore

Dissertations, Master's Theses and Master's Reports

The Nickel based Carbon Monoxide Dehydrogenase (CODH) is an anaerobic metalloenzyme responsible for the reversible conversion of CO and water into CO2 and 2 protons and 2 electrons. This enzyme has importance in the environment as one of Earth’s first carbon fixation pathways, and for human uses as a potential source of biofuels and other commodity chemicals. CODH enzymes are present in a wide array of taxa, many of which are uncultured. In this study we express and purify the catalytic subunit (CooS) of the anaerobic CODH from an uncultured Hydrothermarchaeota JdFR-17 co-expressed with the nickel insertion accessory protein (CooC) …

The Role Of Rad51 In Trichomonas Vaginalis, Dominique Hall

University of the Pacific Theses and Dissertations

Drug resistance to the current treatments on the market is on the rise, therefore there is strong interest in understanding what could be causing the resistance, how resistance could be spreading through the population, and finding some possible new drug targets. One protein of interest is Radiation Sensitive Protein 51 (Rad51). It is a protein that is involved in homologous recombination as well as other processes such as DNA damage repair. While Trichomonas vaginalis traditionally has been known to replicate via binary fission, a modified form of closed mitosis, there is some evidence that meiosis, or at least some form …

Molecular Mechanisms Of Aberrant Protein Glycosylation In Pancreatic Cancer Stemness And Metastasis, Frank Leon

Theses & Dissertations

A myriad of genetic and other abnormal changes underlies the aggressiveness and dissemination properties observed in pancreatic cancer (PC). Aberrant protein glycosylation is a commonly observed feature in PC. The modification of protein O-glycosylation is mediated by glycosyltransferases, which attach and sequentially elongate monosaccharides on Serine/Threonine (Ser/Thr) motifs. Aberrant glycosylation is recognized as an emerging hallmark of cancer where a disruption in normal glycosylation results in irregular O-glycans.

This dissertation research has investigated the consequences of aberrant protein glycosylation on stemness and enhancement of metastatic properties in pancreatic ductal adenocarcinoma (PDAC). Several publications have reported aberrant O-glycosylation increases in oncogenic …

Analysis Of Single-Site Cysteine Mutation, I412c, In Human A Glycine Receptor States To Further Refine Structure And Allostery, Leah Engquist

Honors Program: Senior Projects (Public)

The glycine receptor (GlyR) is the major inhibitory receptor in the brain and spinal cord. A member of the pentameric ligand gated ion channel superfamily, crystal structures are available but there are still unresolved areas, specifically the C-terminal tail and TM3-TM4 intracellular loop. Further refinement can provide deeper understanding of the molecular mechanism and allow the creation of novel therapeutics to modulate its function. We propose to insert a single cysteine mutation, I412C, into a Cys null background (C41S/C290A/C345S) to study non- conducting states (resting, desensitized) or with F207G/A288G mutations to study the open state. Purified, reconstituted GlyR is crosslinked …

High Altitude Ballooning As A Platform For Measuring Ozone Uptake Over Agricultural Landscapes, Thomas Sykora, Mark Potosnak

DePaul Discoveries

Measuring plant health is a key aspect in maximizing crop outputs. One often overlooked risk to crop fields is damage caused by stomatal ozone uptake; measuring this uptake is an important tool in understanding crop losses. Traditional methods for measuring plant ozone uptake are prohibitively expensive and rely on equipment that cannot easily be moved. Here, we propose high-altitude weather ballooning as a cost-effective alternative for measuring ozone uptake on a regional (~10 km) spatial scale. Ozonesounde data was obtained with weather balloons launched from the National Oceanic and Atmospheric Administration research station in Boulder, Colorado. This data was then …

Mucinomics: A Bioinformatic Analysis Of Snail Mucins, And Their Function, Maxwell B. Mcdermott

Theses and Dissertations

This thesis outlines the current research on secreted snail mucus, highlighting the potential of this biopolymer, and also demonstrates a research strategy to fulfill the unmet need of examining the hierarchical structures that lead to the enormous biological and chemical diversity of snail mucus genes.

Development Of High Value Oil Traits Using The Model Oilseed Crop Camelina Sativa, Evan Updike

Department of Biochemistry: Dissertations, Theses, and Student Research

Plant oils are an important source of food, fuel, and feed in our society today. The oil found in the seeds of plants is composed almost entirely of triacylglycerol (TAG) molecules, which consist of three fatty acids esterified to a glycerol backbone. As crude oil supplies decline, vegetable oils are gaining traction as a renewable substitute to petroleum-based materials in fuels, lubricants, and specialty oleochemicals. However, as it currently stands vegetable oils do not possess the properties necessary to fill the void of a petroleum free world.

To address this problem, plant biotechnologists have done extensive work on genetic engineering …

Determination Of The Structure, Function, And Mechanism Of Type Iv Crispr-Cas Prokaryotic Defense Systems, Hannah Nicole Taylor

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Bacteria are under constant threat of invasion by bacteriophage (viruses which infect bacteria). To prevent bacteriophage from entering and overtaking the bacteria, bacteria utilize defense systems to identify and destroy foreign elements. One method of defense is called CRISPR-Cas (Clustered Regularly Interspaced Short Palindromic Repeats – CRISPR-Associated). Many different bacteria and most archaea use CRISPR-Cas systems. There are many diverse types of CRISPR-Cas systems, each of which provides defense in a slightly different way. One such CRISPR-Cas type is called type IV. The type IV CRISPR-Cas system is poorly understood and there are very few studies published on type IV …

Ahr Expression On Rorc-Expressing Immune Cells Is Essential For I3c-Mediated Protection Against Colitis, Michal C. Williams

Senior Theses

Colitis is an inflammatory bowel disorder (IBD) whose etiology is attributed to modification in the luminal microbiota and dysregulation in the immune response. Indole is a signaling molecule which is naturally produced by gut luminal microbiota. Indole-3- carbinol (I3C) is a compound commonly found in vegetables and a ligand for the aryl hydrocarbon receptor (AhR). Previous studies have detected decreased expression and activation on the AhR receptor in colitis patients, thought to possibly alter gut microbiota metabolism, subsequently promoting colitis. 1 AhR, expressed in a variety of immune and epithelial cells, contributes to gut homeostasis by affecting vital mediators such …

Investigating The Mechanism Of The Escherichia Coli Atp-Binding Cassette (Abc) Transporter Metni, Matthew Foronda

Master's Theses

Chemical homeostasis is a baseline requirement for any cell to survive. ATP-binding cassette (ABC) transporters play a vital role in homeostasis by importing nutrients and exporting toxins against their concentration gradients by utilizing the energy of ATP hydrolysis. Malfunctioning ABC transporters cause a variety of health problems, including cystic fibrosis, Stargardt’s disease (vision loss), and the development of drug-resistant tumors. An important step in solving these medical issues is to first understand the structure and mechanism of ABC transporters. Various studies have made great strides in depicting the structure and details of different ABC transporters and their mechanisms, however, many …