Biochemistry, Biophysics, and Structural Biology Commons^™

Evolutionary Convergence Of The Caffeine Biosynthetic Pathway In Chocolate Followed Duplication Of A Constrained Ancestral Enzyme, Andrew J. O'Donnell

Masters Theses

Caffeine biosynthesis is widely distributed in flowering plants and requires three consecutive methylation steps of xanthine alkaloids. Genes that have previously been reported to participate in the multi-step pathway in Coffea sp. (coffee) and Camellia sinensis (tea) encode members of the SABATH family of methyltransferases. Two genes highly expressed in fruits of Theobroma cacao (cacao) are orthologous to the caffeine genes in tea and appear to have diversified following gene duplication. Biochemical characterization of the enzymes (XMTs) encoded by these genes strongly suggest an unprecedented major pathway to theobromine, a precursor to caffeine. These findings imply that caffeine biosynthesis evolved …

Microbial Community Analysis From Petroleum-Contaminated Sediments With High Magnetic Susceptibility, Anja E. Williams

Masters Theses

Microbial biodegradation is an increasingly incorporated method of hydrocarbon mineralization during oil spill cleanup. To monitor biodegradation, magnetic susceptibility (MS) is a geophysical tool that detects changes in sediments’ magnetism, possibly caused by microbial activity, for example by anaerobic iron reduction that can induce magnetite formation. Furthermore, the high input of carbon from an oil spill makes other required nutrients, like nitrogen, limiting. The first study goal was to characterize the microbial communities present at high levels of MS with 16S rRNA gene clone libraries at the petroleum-contaminated site in Bemidji, Minnesota. Sequences with high similarity to known iron-reducers were …

Curcumin: A Multi-Dimensional Approach To Pancreatic Cancer Targeting Cell Death And Exosomes, Carlos J. Diaz Osterman

Loma Linda University Electronic Theses, Dissertations & Projects

Pancreatic cancer is currently one of the most difficult diseases to treat due to difficulty of detection and the aggressive nature of the disease. In addition, pancreatic cancer has the highest mortality rates compared to other cancer types. These mortality rates are attributable in part to increasing resistance to cancer therapy. Cancer therapy resistance is caused by adaptations that favor survival within cancer cells and their environment, termed the tumor microenvironment. Intracellular adaptations include the overexpression of resistance-linked genes, such as the inhibitor of apoptosis (IAP) family of proteins and overall resistance to cell death. Adaptations in the tumor microenvironment …

Crotalus Snake Venom Preconditioning To Prevent Surgical Brain Injury, Cherine Hee-Sun Kim

Loma Linda University Electronic Theses, Dissertations & Projects

Preventive measures are increasingly relevant to medical practice. Preconditioning, a preemptive therapy that administers mildly harmful stimuli to induce endogenous protective mechanisms before major injury, has been shown to minimize injury in many animal models. Given the elective nature of most neurosurgical procedures, the surgical brain injury (SBI) rodent model provides an ideal platform for preconditioning. Our work shows that preconditioning with Crotalus rattlesnake venom, known for its hemorrhagic and inflammatory effects, mitigates some harmful effects of SBI. We have identified two proteins of interest in Crotalus venom: snake venom metalloproteinase (SVMP), an enzyme with hemorrhagic effects, and phospholipase A2 …

A Model Of How Different Biology Experts Explain Molecular And Cellular Mechanisms, Caleb M. Trujillo, Trevor R. Anderson, Nancy J. Pelaez

PIBERG Publications

Constructing explanations is an essential skill for all science learners. The goal of this project was to model the key components of expert explanation of molecular and cellular mechanisms. As such, we asked: What is an appropriate model of the components of explanation used by biology experts to explain molecular and cellular mechanisms? Do explanations made by experts from different biology subdisciplines at a university support the validity of this model? Guided by the modeling framework of R. S. Justi and J. K. Gilbert, the validity of an initial model was tested by asking seven biologists to explain a molecular …

A Study Of Allylic Aminations As Catalyzed By Heterobimetallic Pd-Ti Complexes, Diana Lauren Ellis

Theses and Dissertations

Heterobimetallic complexes present a unique approach to catalyzing challenging reactions. By having two metals in close proximity to each other, the metals are able to interact and alter their electronics in a way that simple organic ligands (carbon, nitrogen, sulfur etc.) cannot. Our studies of heterobimetallic complexes focus on a Pd–Ti complex. The complex features a dative interaction between the palladium and the titanium held together by a phosphonamide scaffold. This interaction increases the electrophilicity of the palladium and makes it a very suitable catalyst for allylic amination reactions. We have conducted extensive studies of this catalyst in allylic aminations, …

Isquest: Finding Insertion Sequences In Prokaryotic Sequence Fragment Data, Abhishek Biswas, David T. Gauthier, Desh Ranjan, Mohammad Zubair

Computer Science Faculty Publications

Motivation: Insertion sequences (ISs) are transposable elements present in most bacterial and archaeal genomes that play an important role in genomic evolution. The increasing availability of sequenced prokaryotic genomes offers the opportunity to study ISs comprehensively, but development of efficient and accurate tools is required for discovery and annotation. Additionally, prokaryotic genomes are frequently deposited as incomplete, or draft stage because of the substantial cost and effort required to finish genome assembly projects. Development of methods to identify IS directly from raw sequence reads or draft genomes are therefore desirable. Software tools such as Optimized Annotation System for Insertion Sequences …

Chronic Inflammation As A Result Of Hepatitis C Virus Infection: A Review Of The Literature, Samantha L. Lane

DePaul Discoveries

Approximately 170 million people are infected with Hepatitis C virus (HCV) worldwide^5,6. It is estimated that roughly 80% of those infected suffer from persistent infection with the virus; this persistence of infection is progressive, and over time can lead to fibrosis, cirrhosis, and hepatocellular carcinoma⁷. Chronic inflammation and apoptotic deregulation are both hallmarks of chronic HCV infection, and many molecular pathways are initiated in both the innate and adaptive immune responses during infection with this viral pathogen. The aim of this review was to survey some of the major molecular mechanisms responsible for the induction of …

Proton Pumping In Cytochrome C Oxidase, Jianxun Lu

Dissertations, Theses, and Capstone Projects

Cytochrome c oxidase (CcO) is a large trans-membrane protein, which is the final enzyme in the respiratory electron transport chain in mitochondria or aerobic bacteria. It implements proton pumping through the mitochondrial membrane against the electrochemical gradient, by utilizing the chemical energy released by reducing O₂ to water. The active site of the chemical reaction is called the Binuclear Center (BNC) that is made up of heme a₃, Cu_B, a Tyrosine residue and their ligands. The protein is reduced four times by electron from cytochromes c to reduce O₂ and to generate four different …

The Effects Of Sustained, High-Velocity Exercise On Gene Expression In California Yellowtail (Seriola Lalandi), Kelli Hatter

Undergraduate Honors Theses

California Yellowtail muscle fibers have been observed to exhibit two drastically different development patterns resulting from the speeds at which they are exercised. When fish are exercised at a moderate rate their epaxial fast-twitch muscle fibers grow in diameter—hypertrophy; when they are exercised at a fast speed, more new epaxial fast-twitch muscle fibers are produced—hyperplasia. To determine the underlying reason for this difference in muscle development, my summer research project and honors thesis exercised fish at: fast, moderate, and control speeds for a sustained amount of time to determine what is happening on a cellular level to cause the observed …

Genomic And Experimental Evidence For Multiple Metabolic Functions In The Rida/Yjgf/Yer057c/Uk114 (Rid) Protein Family, Thomas D. Niehaus, Svetlana Gerdes, Kelsey Hodge-Hanson, Aleksey Zhukov, Arthur J L Cooper, Mona Elbadawi-Sidhu

NYMC Faculty Publications

BACKGROUND: It is now recognized that enzymatic or chemical side-reactions can convert normal metabolites to useless or toxic ones and that a suite of enzymes exists to mitigate such metabolite damage. Examples are the reactive imine/enamine intermediates produced by threonine dehydratase, which damage the pyridoxal 5'-phosphate cofactor of various enzymes causing inactivation. This damage is pre-empted by RidA proteins, which hydrolyze the imines before they do harm. RidA proteins belong to the YjgF/YER057c/UK114 family (here renamed the Rid family). Most other members of this diverse and ubiquitous family lack defined functions.

RESULTS: Phylogenetic analysis divided the Rid family into a …

Pqqd Is A Novel Peptide Chaperone That Forms A Ternary Complex With The Radical S-Adenosylmethionine Protein Pqqe In The Pyrroloquinoline Quinone Biosynthetic Pathway, John A. Latham, Anthony T. Iavarone, Ian Barr, Prerak V. Juthani, Judith P. Klinman

Natural Sciences and Mathematics | Faculty Scholarship

Pyrroloquinoline quinone (PQQ) is a product of a ribosomally synthesized and post-translationally modified pathway consisting of five conserved genes, pqqA-E. PqqE is a radical S-adenosylmethionine (RS) protein with a C-terminal SPASM domain, and is proposed to catalyze the formation of a carbon-carbon bond between the glutamate and tyrosine side chains of the peptide substrate PqqA. PqqD is a 10-kDa protein with an unknown function, but is essential for PQQ production. Recently, in Klebsiella pneumoniae (Kp), PqqD and PqqE were shown to interact; however, the stoichiometry and KD were not obtained. Here, we show that the PqqE and PqqD interaction transcends …

Pharmacology Of Organic Cation Transporters: Focus On Structure-Function Relationships In Oct3 (Slc22a3), Dan C. Li

Undergraduate Theses—Unrestricted

Organic Cation Transporters (OCTs) are polyspecific, facilitative transporters that play major roles in metabolite and drug clearance. OCTs are promising drug targets and elucidating their mechanisms of substrate recognition is crucial for rational drug design. OCT-mediated transport of polyvalent cations remains unexplored. OCT-expressing Xenopus laevis oocytes were used to assess transport of polyamines, ubiquitous polyvalent cations of broad physiological import, but for which transport mechanisms are unknown. Dose-response analysis of radiolabelled substrate uptake revealed that polyamines are relatively low affinity, but high turnover substrates for OCTs compared to model substrate methyl-4-phenylpyridinium (MPP⁺). Polyamine analogs of varying hydrophobic character …

Preventing Farnesylation Of The Dynein Adaptor Spindly Contributes To The Mitotic Defects Caused By Farnesyltransferase Inhibitors, Andrew J. Holland, Rita M. Reis, Sherry Niessen, Cláudia Pereira, Douglas A. Andres, H. Peter Spielmann, Don W. Cleveland, Arshad Desai, Reto Gassmann

Molecular and Cellular Biochemistry Faculty Publications

The clinical interest in farnesyltransferase inhibitors (FTIs) makes it important to understand how these compounds affect cellular processes involving farnesylated proteins. Mitotic abnormalities observed after treatment with FTIs have so far been attributed to defects in the farnesylation of the outer kinetochore proteins CENP-E and CENP-F, which are involved in chromosome congression and spindle assembly checkpoint signaling. Here we identify the cytoplasmic dynein adaptor Spindly as an additional component of the outer kinetochore that is modified by farnesyltransferase (FTase). We show that farnesylation of Spindly is essential for its localization, and thus for the proper localization of dynein and its …

Alternate Formulations For Optimization Of Plga Microsphere-Comprised Glucose Biosensor Coating Against Foreign Body Inflammation., Klair Lubonja

Honors Scholar Theses

In this study, dexamethasone loaded PLGA microsphere/PVA hydrogel composites were explored as an outer drug-eluting coating for implantable biosensors to provide defense against acute inflammation of the foreign body response. The microspheres, with a dexamethasone release target period of approximately two weeks, were manufactured under various conditions: different co-solvents and homogenization speeds. Particle size measurement, scanning electron microscopy and differential scanning calorimetry were performed for all the microsphere formulations prepared. The addition of acetone as a co-solvent for dexamethasone, at normal homogenization speeds, shows promise for the improvement of drug loading and the attainment of superior release profiles. PLGA microsphere/PVA …

Biological Significance Of Photoreceptor Photocycle Length: Vivid Photocycle Governs The Dynamic Vivid-White Collar Complex Pool Mediating Photo-Adaptation And Response To Changes In Light Intensity, Arko Dasgupta, Chen-Hui Chen, Changhwan Lee, Amy S. Gladfelter, Jay C. Dunlap, Jennifer J. Loros

Dartmouth Scholarship

Most organisms on earth sense light through the use of chromophore-bearing photoreceptive proteins with distinct and characteristic photocycle lengths, yet the biological significance of this adduct decay length is neither understood nor has been tested. In the filamentous fungus Neurospora crassa VIVID (VVD) is a critical player in the process of photoadaptation, the attenuation of light-induced responses and the ability to maintain photosensitivity in response to changing light intensities. Detailed in vitro analysis of the photochemistry of the blue light sensing, FAD binding, LOV domain of VVD has revealed residues around the site of photo-adduct formation that influence the stability …

Three-Dimensional Ideal Gas Reference State Based Energy Function, Avdesh Mishra

LSU New Orleans Theses and Dissertations

Energy functions are found to be a key of protein structure prediction. In this work, we propose a novel 3-dimensional energy function based on hydrophobic-hydrophilic properties of amino acid where we consider at least three different possible interaction of amino acid in a 3-dimensional sphere categorized as hydrophilic versus hydrophilic, hydrophobic versus hydrophobic and hydrophobic versus hydrophilic. Each of these interactions are governed by a 3-dimensional parameter alpha used to model the interaction and 3-dimensional parameter beta used to model weight of contribution. We use Genetic Algorithm (GA) to optimize the value of alpha, beta and Z-score. We obtain three …

Proteasome-Mediated Proteolysis Of The Polyglutamine-Expanded Androgen Receptor Is A Late Event In Spinal And Bulbar Muscular Atrophy (Sbma) Pathogenesis., Erin M Heine, Tamar R Berger, Anna Pluciennik, Christopher R Orr, Lori Zboray, Diane E Merry

Department of Biochemistry and Molecular Biology Faculty Papers

Proteolysis of polyglutamine-expanded proteins is thought to be a required step in the pathogenesis of several neurodegenerative diseases. The accepted view for many polyglutamine proteins is that proteolysis of the mutant protein produces a "toxic fragment" that induces neuronal dysfunction and death in a soluble form; toxicity of the fragment is buffered by its incorporation into amyloid-like inclusions. In contrast to this view, we show that, in the polyglutamine disease spinal and bulbar muscular atrophy, proteolysis of the mutant androgen receptor (AR) is a late event. Immunocytochemical and biochemical analyses revealed that the mutant AR aggregates as a full-length protein, …

Revealing Important Nocturnal And Day-To-Day Variations In Fire Smoke Emissions Through A Multiplatform Inversion, Pablo E. Saide, David A. Peterson, Arlindo Da Silva, Bruce Anderson, Luke D. Ziemba, Glenn Diskin, Glen Sachse, Johnathan Hair, Carolyn Butler, Marta Fenn, Jose L. Jimenez, Pedro Campuzano-Jost, Anne E. Perring, Joshua P. Schwarz, Milos Z. Markovic, Phil Russell, Jens Redemann, Yohei Shinozuka, David G. Streets, Fang Yan, Jack Dibb, Robert Yokelson, O. Brian Toon, Edward Hyer, Gregory R. Carmichael

Chemistry and Biochemistry Faculty Publications

We couple airborne, ground-based, and satellite observations; conduct regional simulations; and develop and apply an inversion technique to constrain hourly smoke emissions from the Rim Fire, the third largest observed in California, USA. Emissions constrainedwithmultiplatform data show notable nocturnal enhancements (sometimes over a factor of 20), correlate better with daily burned area data, and are a factor of 2–4 higher than a priori estimates, highlighting the need for improved characterization of diurnal profiles and day-to-day variability when modeling extreme fires. Constraining only with satellite data results in smaller enhancements mainly due to missing retrievals near the emissions source, suggesting that …

Reverse Gyrase Is Not Necessary For Survival Of Hyperthermophilic Archaeon Pyrococcus Furiosus, Farshid Taghizadeh, Michael S. Bartlett

Student Research Symposium

Reverse gyrase is the only known topoisomerase enzyme with positive supercoiling activity on covalently-closed DNA. This positive supercoiling is required to prevent DNA from denaturation at high temperatures. The gene that codes for this protein is present in all hyperthermophiles and absent from all mesophilic and thermophilic genomes, suggesting that this enzyme is the only hyperthermophile-specific protein. To investigate if this protein is vital for the cells, we knocked out its gene from the genome of living organism Pyrococcus furiosus. Pyrococcus furiosus is a hyperthermophilic archaeon that grows between 70°C to 103°C with an optimum growth temperature of 100°C. …

Probing Buffer-Specific Effects On Nucleotide Binding To Reca Using Difference Fourier Transform Infrared Spectroscopy, Joshua E. Temple

Senior Honors Projects, 2010-2019

The Escherichia coli protein RecA catalyzes the strand exchange reaction used in DNA repair and genetic recombination. RecA is also a target for inhibiting microbial antibiotic resistance, understanding cancer propagation, and characterizing neurodegenerative disorders. Therefore, understanding factors that affect RecA structure and stability is broadly applicable to many fields. Previous studies in our lab have shown buffer-specific changes in RecA stability and unfolding transitions. These studies suggest only minimal buffer-dependent changes in nucleotide binding and secondary structure but do not explain the significant differences in RecA stability and unfolding profiles. Here we have employed various biochemical and spectroscopic techniques to …

Structural Elucidation Of Aggr-Activated Regulator, Aar, In Enteroaggregative Escherichia Coli, Andrew Heindel

Senior Honors Projects, 2010-2019

Travelers’ Diarrhea is the number one cause of childhood death in the world. Enteroaggregative Escherichia coli (EAEC) is one of the main causes of this disease. EAEC adhere to the surface of the intestine and stack in a brick-like pattern. Via an unstudied quorum-sensing mechanism, these bacteria express a variety of virulence factors that lead to diarrhea. The long-term goal of this research is to elucidate the mechanism by which EAEC changes from benign to virulent. A previously-unstudied open-reading frame in EAEC, AggR activated repressor (Aar), has recently been hypothesized to act as one of the major transcription factors influencing …

Biophysical Characterization Of Naturally Occurring Titin M10 Mutations, Michael William Rudloff

Senior Honors Projects, 2010-2019

The giant human muscle proteins titin and obscurin are important for sarcomeric organization, stretch response, and sarcomerogenesis in myofibrils. The extreme C-terminus of titin (the M10 domain) binds to the N-terminus of obscurin (the Ig1 domain) in the M-line, an interaction that is critical for sarcomere stability. The high-resolution structure of human M10 has been solved, along with M10 bound to one of its two known molecular targets, the Ig1 domain of obscurin-like protein. Multiple M10 mutations are linked to limb-girdle muscular dystrophy type 2J (LGMD2J) and tibial muscular dystrophy (TMD), however the effect of the M10 mutations on protein …

Fishing For A Sustainable Future: Aquaponics As A Method Of Food Production, Richard Ramsundar

Student Theses 2015-Present

This thesis compares and explains the advantages aquaponics farming has over modern industrial intensive farming. Through a comparison natural capital usage, conservation, recycling and cost, the thesis advocates for the expansion of aquaponics usage in urban settings. The thesis also explains the history of intensive farming and aquaponics in America, the science of how aquaponics operates, the economic and environmental costs of modern intensive farming versus aquaponics farming, and the social implications of aquaponics. Lastly, I propose a policy that reallocates farm subsidies by modifying the Farm Bill. Then I propose policies that support creating a new standard of farm …

Section Abstracts: Structural Biology, Biochemistry, And Biophysics

Virginia Journal of Science

Abstracts of the Structural Biology, Biochemistry, and Biophysics Section for the 93rd Annual Meeting of the Virginia Academy of Science, May 21-23, 2015, James Madison University, Richmond, Virginia

Section Abstracts: Biology With Microbiology And Molecular Biology

Virginia Journal of Science

Abstracts of the Biology with Microbiology and Molecular Biology Section for the 93rd Annual Meeting of the Virginia Academy of Science, May 21-23, 2015, James Madison University, Richmond, Virginia

An Improved 96-Well Turbidity Assay For T4 Lysozyme Activity, Tasha B. Toro, Thao P. Nguyen, Terry J. Watt

Faculty and Staff Publications

T4 lysozyme (T4L) is an important model system for investigating the relationship between protein structure and function. Despite being extensively studied, a reliable, quantitative activity assay for T4L has not been developed. Here, we present an improved T4L turbidity assay as well as an affinity-based T4L expression and purification protocol. This assay is designed for 96-well format and utilizes conditions amenable for both T4L and other lysozymes. This protocol enables easy, efficient, and quantitative characterization of T4L variants and allows comparison between different lysozymes. Our method: •Is applicable for all lysozymes, with enhanced sensitivity for T4 lysozyme compared to other …

Structural And Functional Interactions Between Bro1 Domain Of Human Alix Protein And Nucleocapsid Packaging Rna Complex From Hiv, Scott Gross

Graduate School of Biomedical Sciences Theses and Dissertations

A virus is only as powerful as its ability to spread. Enveloped retroviruses, namely HIV-1, use exocytosis pathways that normal host cells use to release particles from the plasma membrane. The main pathways of interest in this study are the Endosomal Sorting Complex Required for Transport (ESCRT) and adjacent ALIX pathways. The ESCRT pathway is especially important for degradation of receptor/cargo complexes that form Multi-Vesicular Bodies (MVBs). Currently, there is no known therapy that targets this endosomal pathway, which would prevent the spread of the virus to other cells. The virus has adapted to jump from pathway to pathway when …

Age-Dependent Breakdown Of The Blood Brain Barrier And Associated Changes In S100b Ko Mice, Eric V. Brown

Graduate School of Biomedical Sciences Theses and Dissertations

Autoantibodies play an important role in many autoimmune diseases. Recent research has shown that breakdown of the blood brain barrier (BBB) occurs concomitant to generation of brain reactive autoantibodies in many neurodegenerative diseases, which serve as biomarkers and drivers of pathology. SI00B, a calcium binding protein found most highly expressed in astrocytes which ensheathe the BBB, has many functions in neural development and signaling. Currently literature indicates that S100B KO mice develop normally, with no phenotypic abnormalities. Here, it is demonstrated that S100B KO mice seem to develop a chronic BBB breakdown similar to that seen in human neurodegenerative diseases. …

Evolution Of Sperm Activation In Nematodes, Qing Wei

Graduate School of Biomedical Sciences Theses and Dissertations

Three hermaphroditic species in the nematode genus Caenorhabditis - C. elegans, C. briggsae and C. tropicalis evolved independently from a male/female ancestor. Thus, the hermaphroditic reproductive system provides an ideal subject for studying the origin of new complex traits. Previous studies implied that this transition required two steps: a mutation that caused XY animals to make sperm, and a mutation that allowed XX spermatids to activate and fertilize oocytes. In this dissertation, I show how sperm activation is regulated in different hermaphroditic species.

First, I used TALENs to create genetic tools for C. briggsae and C. tropicalis. These tools were …