Biochemistry Commons^™

Biochemical And Biophysical Studies Of Vision Loss: Cataract And Macular Degeneration, Michael C. Vega

Graduate Research Theses & Dissertations

Age-related biochemical and biophysical changes to the structural properties of ocular lens membrane proteins and retinal pigment epithelium (RPE) cells are thought to play an underlying role in the etiology and progression of age-related cataract and macular degeneration. This dissertation work studies biophysical properties of biomimetic membranes to aid in further understanding of the role of aquaporin-0 in cataract pathogenesis and the biochemical composition of age-related pigmented granules, melanolipofuscin, and their implications in macular degeneration.

Cataract is the leading cause of blindness globally and is characterized by the opacification of the ocular lens. As a key optical component of the …

Thermostability And Excision Activity Of Polymorphic Forms Of Hogg1, Kathryn Mouzakis

Chemistry and Biochemistry Faculty Works

Objectives: Reactive oxygen species (ROS) oxidize guanine residues in DNA to form 7,8-dihydro-oxo-2′-deoxyguanosine (8oxoG) lesions in the genome. Human 8-oxoguanine glycosylase-1 (hOGG1) recognizes and excises this highly mutagenic species when it is base-paired opposite a cytosine. We sought to characterize biochemically several hOGG1 variants that have been found in cancer tissues and cell lines, reasoning that if these variants have reduced repair capabilities, they could lead to an increased chance of mutagenesis and carcinogenesis. Results: We have over-expressed and purified the R46Q, A85S, R154H, and S232T hOGG1 variants and have investigated their repair efficiency and thermostability. The hOGG1 variants showed …

Green Chemistry In United States Science Policy, Emily Jarvis

Chemistry and Biochemistry Faculty Works

Although optimizing each of the Twelve Principles of Green Chemistry is necessary for achieving the greenest possible chemistry in the lab, there are additional creative ways to promote green chemistry not encompassed by these. Communicating to society the importance of green chemistry beyond the laboratory is essential. Of course, doing this effectively presents distinct challenges to those encountered when designing chemical syntheses. Here, I discuss several observations from my time working in federal science policy as an American Association for the Advancement of Science/American Chemical Society Congressional Science Fellow in the United States Senate. I suggest a practical science policy …

Automatic 13C Chemical Shift Reference Correction Of Protein Nmr Spectral Data Using Data Mining And Bayesian Statistical Modeling, Xi Chen

Theses and Dissertations--Molecular and Cellular Biochemistry

Nuclear magnetic resonance (NMR) is a highly versatile analytical technique for studying molecular configuration, conformation, and dynamics, especially of biomacromolecules such as proteins. However, due to the intrinsic properties of NMR experiments, results from the NMR instruments require a refencing step before the down-the-line analysis. Poor chemical shift referencing, especially for ¹³C in protein Nuclear Magnetic Resonance (NMR) experiments, fundamentally limits and even prevents effective study of biomacromolecules via NMR. There is no available method that can rereference carbon chemical shifts from protein NMR without secondary experimental information such as structure or resonance assignment.

To solve this problem, we …

Computational Tools For The Untargeted Assignment Of Ft-Ms Metabolomics Datasets, Joshua Merritt Mitchell

Theses and Dissertations--Molecular and Cellular Biochemistry

Metabolomics is the study of metabolomes, the sets of metabolites observed in living systems. Metabolism interconverts these metabolites to provide the molecules and energy necessary for life processes. Many disease processes, including cancer, have a significant metabolic component that manifests as differences in what metabolites are present and in what quantities they are produced and utilized. Thus, using metabolomics, differences between metabolomes in disease and non-disease states can be detected and these differences improve our understanding of disease processes at the molecular level. Despite the potential benefits of metabolomics, the comprehensive investigation of metabolomes remains difficult.

A popular analytical technique …

Assessing The Effects Of Antioxidants On Oxidative Stress In Triple-Negative Breast Cancer Cells Via Protein Carbonylation, Dalton Sheffield

Honors Theses

Breast cancer is a highly complex, heterogeneous disease, and potential effective treatment options are being continuously researched in order to provide a method of halting cancer growth and metastasis. One area in particular involves alleviating oxidative stress within cancer cells. Oxidative stress is generally defined as an imbalance between oxidant and antioxidant species in favor of the oxidants. Cancer cells use this oxidative stress to initiate carcinogenesis in cells through different mechanisms, and protein carbonylation is a primary biomarker used to quantify oxidative stress levels. Antioxidant compounds are those used to alleviate oxidative stress within cells. Here breast cancer cells …

The Role Of Irf1 In The Brain And In Adaptive Responses Of Astrocytes, Andrew Hoskins

Theses and Dissertations

In neurodegenerative diseases, the CNS becomes inflamed through activation of pathways, including the NF-B pathway. Some of the therapies for those diseases target neuroinflammatory pathways. Here, we explore the mechanisms for the upregulation of a subset of genes following a restimulation of the NF-B pathway. We discover that this upregulation occurs independent of IRF1 expression and type 1 interferon signaling. A knockdown of IRF1 using siRNA and an inhibition of JAK proteins using inhibitor AG490 both had no effect on priming. A secreted factor was found to upregulate the expression of both this subset of genes and genes encoding pro-inflammatory …

Cellular Mechanisms By Which Alcohol Promotes Hiv Protease Inhibitor-Induced Hepatotoxicity, Michael Hinton

Theses and Dissertations

CELLULAR MECHANISMS BY WHICH ALCOHOL PROMOTES HIV PROTEASE INHIBITOR-INDUCED HEPATOTOXICITY

Michael Hinton, B.S.

A dissertation submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy at Virginia Commonwealth University

Virginia Commonwealth University, 2019

Major Director: Huiping Zhou

Professor, Department of Microbiology and Immunology

The development of highly-active-antiretroviral therapy(HAART) has allowed management of HIV and extended the lives of those infected. Alcohol abuse, which is very common in HIV-1 infected patients, is one of the most important co-morbid risk factors for liver injury and has been associated with the occurrence of serious metabolic syndrome and subsequent discontinuation …

Near-Infrared Fluorescent Probes For Sensitive Determination Of Lysosomal & Mitochondrial Ph In Live Cells, Wafa Mazi

Dissertations, Master's Theses and Master's Reports

Varied intracellular pH levels are critical for various physiological processes such as enzymatic activity, cell proliferation and apoptosis, ion transport, and muscle contraction. Cellular compartments, like lysosomes, must retain an acidic environment (pH ~ 4.5) to activate hydrolytic enzymes necessary for the breakdown of large biomolecules. Another cellular organelle, the mitochondria, provides the cell with energy and must retain an alkalis environment (pH ~ 8.0) for proper function. Substantial lysosomal and mitochondrial pH deviation is associated with cellular dysfunction and disease. Therefore, the precise detection of lysosomal and mitochondrial pH is essential to provide a better understanding of cellular physiological …

The Functional Role Of Rna Binding Protein Rbms3 As A Tumor Promoter In Triple-Negative Breast Cancer Cells, Yuting Zhou

Theses and Dissertations--Molecular and Cellular Biochemistry

RBMS3 belongs to the family of c-myc gene single-strand binding proteins (MSSPs) that play important roles in transcriptional regulation. Here, we show that RBMS3 functions as a tumor promoter in triple-negative breast cancer (TNBC), a highly aggressive BC subtype. Analysis of RBMS3 expression shows that RBMS3 is upregulated at both mRNA and protein levels in TNBC cells. Functionally, overexpression of RBMS3 increases cell migration, invasion and cancer stem cell (CSC) behaviors. Moreover, RBMS3 induces expression of epithelial-mesenchymal transition (EMT) and CSC markers. Conversely, loss of RBMS3 in TNBC BT549 cells inhibits cell proliferation, migration and mesenchymal phenotype. Correlation analysis shows …

Biochemical Approaches For The Diagnosis And Treatment Of Lafora Disease, Mary Kathryn Brewer

Theses and Dissertations--Molecular and Cellular Biochemistry

Glycogen is the sole carbohydrate storage molecule found in mammalian cells and plays an important role in cellular metabolism in nearly all tissues, including the brain. Defects in glycogen metabolism underlie the glycogen storage diseases (GSDs), genetic disorders with variable clinical phenotypes depending on the mutation type and affected gene(s). Lafora disease (LD) is a fatal form of progressive myoclonus epilepsy and a non-classical GSD. LD typically manifests in adolescence with tonic-clonic seizures, myoclonus, and a rapid, insidious progression. Patients experience increasingly severe and frequent epileptic episodes, loss of speech and muscular control, disinhibited dementia, and severe cognitive decline; death …

Investigating Potential Multi-Protein Complexes Involving Emx2, Nicole Enger

All Master's Theses

The neocortex of the mammalian brain is allocated into specialized areas during embryonic development. In mice, as well as humans, the neocortex develops four primary areas: somatosensory, auditory, visual, and motor. In the adult neocortex, the boundaries between these four areas are marked by differences in cellular architecture and gene expression patterns. However, these physical boundaries are not evident during embryonic development of this structure. Rather, the neocortex initially appears uniform across its expanse, with one exception: a handful of proteins that regulate neocortical arealization are generated in gradients across the expanse of the neocortex and control the size of …

Visualizing The Invisible: A Guide To Designing, Printing, And Incorporating Dynamic 3d Molecular Models To Teach Structure–Function Relationships, Michelle Howell, Karin Van Dijk, Christine S. Booth, Tomáš Helikar, Brain A. Couch, Rebecca Roston

Department of Biochemistry: Faculty Publications

Understanding the intricate relationship between macromolecular structure and function represents a central goal of undergraduate biology education (1–3). In teaching complex three-dimensional (3D) concepts, instructors typically depend on static two-dimensional (2D) textbook images or computer-based visualization software, which can lead to unintended misconceptions (4–6). While chemical and molecular kits exist, these models cannot handle the size and detail of macromolecules. Consequently, students may graduate in the life sciences without understanding how structure underlies function or acquiring skills to translate between 2D and 3D molecular models (5, 7). Building on recent technological advances, 3D printing (3DP) potentiates an era in which …

Autophagy In Adipocyte Browning: Emerging Drug Target For Intervention In Obesity, Seung-Hyun Ro, Yura Jang, Jiyoung Bae, Isaac M. Kim, Cameron Schaecher, Zachery D. Shomo

Department of Biochemistry: Faculty Publications

Autophagy, lipophagy, and mitophagy are considered to be the major recycling processes for protein aggregates, excess fat, and damaged mitochondria in adipose tissues in response to nutrient status-associated stress, oxidative stress, and genotoxic stress in the human body. Obesity with increased body weight is often associated with white adipose tissue (WAT) hypertrophy and hyperplasia and/or beige/brown adipose tissue atrophy and aplasia, which significantly contribute to the imbalance in lipid metabolism, adipocytokine secretion, free fatty acid release, and mitochondria function. In recent studies, hyperactive autophagy in WAT was observed in obese and diabetic patients, and inhibition of adipose autophagy through targeted …

Dna Uptake By Type Iv Filaments, Kurt H. Piepenbrink

Department of Biochemistry: Faculty Publications

Bacterial uptake of DNA through type IV filaments is an essential component of natural competence in numerous gram-positive and gram-negative species. Recent advances in the field have broadened our understanding of the structures used to take up extracellular DNA. Here, we review seminal experiments in the literature describing DNA binding by type IV pili, competence pili and the flp pili of Micrococcus luteus; collectively referred to here as type IV filaments. We compare the current state of the field on mechanisms of DNA uptake for these three appendage systems and describe the current mechanistic understanding of both DNA-binding and …

Student Understanding Of Dna Structure–Function Relationships Improves From Using 3d Learning Modules With Dynamic 3d Printed Models, Michelle E. Howell, Christine S. Booth, Sharmin M. Sikich, Tomáš Helikar, Rebecca Roston, Brain A. Couch, Karin Van Dijk

Department of Biochemistry: Faculty Publications

Understanding the relationship between molecular structure and function represents an important goal of undergraduate life sciences. Although evidence suggests that handling physical models supports gains in student understanding of structure–function relationships, such models have not been widely implemented in biochemistry classrooms. Three-dimensional (3D) printing represents an emerging cost-effective means of producing molecular models to help students investigate structure–function concepts. We developed three interactive learning modules with dynamic 3D printed models to help biochemistry students visualize biomolecular structures and address particular misconceptions. These modules targeted specific learning objectives related to DNA and RNA structure, transcription factor-DNA interactions, and DNA supercoiling dynamics. …

Winter Hardiness Of Miscanthus (Iii): Genome‐Wide Association And Genomic Prediction For Overwintering Ability In Miscanthus Sinensis, Hongxu Dong, Lindsay V. Clark, Alexander E. Lipka, Joe E. Brummer, Katarzyna Glowacka, Megan C. Hall, Kweon Heo, Xiaoli Jin, Junhua Peng, Toshihiko Yamada, Bimal Kumar Ghimire, Ji Hye Yoo, Chang Yeon Yu, Hua Zhao, Stephen P. Long, Erik J. Sacks

Department of Biochemistry: Faculty Publications

Overwintering ability is an important selection criterion for Miscanthus breeding in temperate regions. Insufficient overwintering ability of the currently leading Miscanthus biomass cultivar, M. ×giganteus (M×g) ‘1993–1780’, in regions where average annual minimum temperatures are −26.1°C (USDA hardiness zone 5) or lower poses a pressing need to develop new cultivars with superior cold tolerance. To facilitate breeding of Miscanthus, this study characterized phenotypic and genetic variation of overwintering ability in an M. sinensis germplasm panel consisting of 564 accessions, evaluated in field trials at three locations in North America and two in Asia. Genome‐wide association (GWA) and genomic …

Chronic Alcohol Exposure Alters Circulating Insulin And Ghrelin Levels: Role Of Ghrelin In Hepatic Steatosis, Karuna Rasineni, Paul G. Thomes, Jacy L. Kubik, Edward N. Harris, Kusum K. Kharbanda, Carol A. Casey

Department of Biochemistry: Faculty Publications

Fatty liver is the earliest response of the liver to excessive ethanol consumption. Central in the development of alcoholic steatosis is increased mobilization of nonesterified free fatty acids (NEFAs) to the liver from the adipose tissue. In this study, we hypothesized that ethanol-induced increase in ghrelin by impairing insulin secretion, could be responsible for the altered lipid metabolism observed in adipose and liver tissue. Male Wistar rats were fed for 5–8 wk with control or ethanol Lieber-DeCarli diet, followed by biochemical analyses in serum and liver tissues. In addition, in vitro studies were conducted on pancreatic islets isolated from experimental …

The Mitochondrial Nad+ Transporter (Ndt1) Plays Important Roles In Cellular Nad+ Homeostasis In Arabidopsis Thaliana, Izabel De Souza Chaves, Elias Feitosa-Araujo, Alexandra Florian, David B. Medeiros, Paula Da Fonseca-Pereira, Lennart Charton, Elmien Heyneke, Jorge A. C. Apfata, Marcel V. Pires, Tabea Mettler-Altmann, Wagner L. Arajuo, H. Ekkehard Neuhaus, Ferdinando Palmieri, Toshihiro Obata, Andreas P. M. Weber, Nicole Linka, Alisdair R. Femie, Adriano Nunes-Nesi

Department of Biochemistry: Faculty Publications

Nicotinamide adenine dinucleotide (NAD⁺) is an essential coenzyme required for all living organisms. In eukaryotic cells, the final step of NAD⁺ biosynthesis is exclusively cytosolic. Hence, NAD⁺ must be imported into organelles to support their metabolic functions. Three NAD⁺ transporters belonging to the mitochondrial carrier family (MCF) have been biochemically characterized in plants. AtNDT1 (At2g47490), focus of the current study, AtNDT2 (At1g25380), targeted to the inner mitochondrial membrane, and AtPXN (At2g39970), located in the peroxisomal membrane. Although AtNDT1 was presumed to reside in the chloroplast …

Acyltransferases And Methods Of Using, Edgar Cahoon, Umidjon Iskandarov, Hae Jin Kim, Jillian Collins-Silva

Department of Biochemistry: Faculty Publications

Provided herein are novel acyltransferases and methods of using such novel acyltransferases in making medium - chain fatty acids.

Charting A New Frontier Of Science By Integrating Mathematical Modeling To Understand And Predict Complex Biological Systems, Katharine White, Kira Mcentire, Nicole R. Buan, Kingshuk Ghosh, Lecia Robinson, Elisar Barbar

Department of Biochemistry: Faculty Publications

Biological systems are staggeringly complex. To untangle this complexity and make predictions about biological systems is a continuous goal of biological research. One approach to achieve these goals is to emphasize the use of quantitative measures of biological processes. Advances in quantitative biology data collection and analysis across scales (molecular, cellular, organismal, ecological) has transformed how we understand, categorize, and predict complex biological systems. Simultaneously, thanks to increased computational power, mathematicians, engineers and physical scientists -- collectively termed theoreticians -- have developed sophisticated models of biological systems at different scales. But there is still a disconnect between the two fields. …

Fatty Acid Transporter 2 Reprograms Neutrophils In Cancer, Filippo Veglia, Vladimir A. Tyurin, Maria Blasi, Alessandra De Leo, Andrew Kossenkov, Laxminarasimha Donthireddy, To Tsun Ki Jerrick, Zach Schug, Subhasree Basu, Fang Wang, Emanuela Ricciotti, Concetta C. Dirusso, Maureen E. Murphy, Robert H. Vonderheide, Paul M. Lieberman, Charles Mulligan, Brian Nam, Neil Hockstein, Gregory Masters, Michael Guarino, Cindy Lin, Yulia Nefedova, Paul N. Black, Valerian E, Kagan, Dmitry Gabrilovich

Department of Biochemistry: Faculty Publications

Polymorphonuclear myeloid derived suppressor cells (PMN-MDSC) are pathologically activated neutrophils that are critically important for the regulation of immune responses in cancer. They contribute to the failure of cancer therapies and are associated with poor clinical outcomes. Despite the recent advances in understanding of the PMN-MDSC biology, the mechanisms responsible for pathological activation of neutrophils are not well defined, which limits selective targeting of these cells. Here, we report that mouse and human PMN-MDSC exclusively up-regulate fatty acid transporter protein 2 (FATP2). Over-expression of FATP2 in PMN-MDSC was controlled by GM-CSF, through the activation of STAT5 transcription factor. Deletion of …

Arsenite Exposure Suppresses Adipogenesis, Mitochondrial Biogenesis And Thermogenesis Via Autophagy Inhibition In Brown Adipose Tissue, Jiyoung Bae, Yura Jang, Heejeong Kim, Kalika Mahato, Cameron Schaecher, Issac M. Kim, Eunju Kim, Seung-Hyun Ro

Department of Biochemistry: Faculty Publications

Arsenite, a trivalent form of arsenic, is an element that occurs naturally in the environment. Humans are exposed to high dose of arsenite through consuming arsenite-contaminated drinking water and food, and the arsenite can accumulate in the human tissues. Arsenite induces oxidative stress, which is linked to metabolic disorders such as obesity and diabetes. Brown adipocytes dissipating energy as heat have emerging roles for obesity treatment and prevention. therefore, understanding the pathophysiological role of brown adipocytes can provide effective strategies delineating the link between arsenite exposure and metabolic disorders. Our study revealed that arsenite significantly reduced differentiation of murine brown …

The Structure Of Pila From Acinetobacter Baumannii Ab5075 Suggests A Mechanism For Functional Specialization In Acinetobacter Type Iv Pili, Leslie A. Ronish, Erik Lillehoj, James K. Fields, Eric J. Sundberg, Kurt Piepenbrink

Department of Biochemistry: Faculty Publications

Type IV pili (T4P) are bacterial appendages composed of protein subunits, called pilins, noncovalently assembled into helical fibers. T4P are essential, in many bacterial species, for processes as diverse as twitching motility, natural competence, biofilm or microcolony formation, and host cell adhesion. The genes encoding type IV pili are found universally in the Gram-negative, aerobic, nonflagellated, and pathogenic coccobacillus Acinetobacter baumannii, but there is considerable variation in PilA, the major protein subunit, both in amino acid sequence and in glycosylation patterns. Here we report the X-ray crystal structure of PilA from AB5075, a recently characterized, highly virulent isolate, at …

The Effects Of Ethynodiol Diacetate On Epinephrine-Treated Glial-Derived Cells, Jesse D. Hunt

Theses and Dissertations

Neuroprotective agents, including specific hormones, may play a role in preventing cellular damage following an ischemic event or brain injury. Astrocytes are specialized glial cells within the central nervous system that play roles in synapse development, blood flow regulation, extracellular homeostasis, synapse function, energy and metabolism (Sofroniew and Vinters, 2009). Research using astrocytes in cell viability studies can be beneficial in deepening our understanding of specific neuroprotective effects of various steroid hormones including progesterone. Progesterone has been shown to have neuroprotective capabilities in the central nervous system (Scarpin et ah, 2009). Although progesterone shows great therapeutic potential, effects of the …

The Involvement Of Progesterone In Glial Cell Viability, Tracee L. Guthrie

Theses and Dissertations

We studied the effects of ethynodiol diacetate on the viability of hydrogen peroxide stressed glial cells. We wanted to determine the extent in which varying concentration of ethynodiol diacetate affect cell viability of cultured glial cells from hydrogen peroxide toxicity. Based on previous studies, we predicted that the viability of hydrogen peroxide-stressed cells will increase with the presence of the progesterone, ethynodiol diacetate, and that cell viability will increase with increasing concentration of ethynodiol diacetate. Glial cells of 1x10° cells/well were introduced into a 96-well plate and allowed to grow for 24 hours in an incubator at 37°C and 5% …

From The Laboratory To The Vineyard—Evolution Of The Measurement Of Grape Composition Using Nir Spectroscopy Towards High-Throughput Analysis, Aoife Power, Vi Khanh Truong, James Chapman, Daniel Cozzolino

Articles

Compared to traditional laboratory methods, spectroscopic techniques (e.g., near infrared, hyperspectralimaging)provideanalystswithaninnovativeandimprovedunderstandingofcomplex issuesbydeterminingseveralchemicalcompoundsandmetabolitesatonce,allowingforthecollection of the sample “fingerprint”. These techniques have the potential to deliver high-throughput options for the analysis of the chemical composition of grapes in the laboratory, the vineyard and before or during harvest, to provide better insights of the chemistry, nutrition and physiology of grapes. Faster computers, the development of software and portable easy to use spectrophotometers and data analytical methods allow for the development of innovative applications of these techniques for the analyses of grape composition.

Mapping The Effects Of 5’ Untranslated Region Deletions On Translation Of 3a In Bmv Rna3, Courtney Marie Bozman

Graduate Research Theses & Dissertations

Brome mosaic virus (BMV) is a positive sense, single-stranded RNA virus that infects a wide array of plant hosts globally including: wheat, oats, maize, barley, and rye. Since its discovery in the 1940s, BMV has served as a model organism in uncovering the modalities of viral infectivity, composition, translation, and replication. In many viruses, it is typically imperative to recruit cellular machinery to produce viral proteins as their coding capacity is limited. Therefore, there are sequences and structures within the leader sequences allow them to be competitive with the eukaryotic mRNAs. Although BMV has been thoroughly researched in these other …

Regulator Of G Protein Signaling-12 (Rgs12) In Dopaminergic And Kappa Opioid Receptor-Dependent Signaling And Behavior, Joshua David Gross

Graduate Theses, Dissertations, and Problem Reports (ETD)

Dopaminergic neurotransmission is critically involved in the etiology and treatment of many psychiatric and neurological disorders. One modulator of dopaminergic neurotransmission is the kappa opioid receptor (KOR) -- a G protein-coupled receptor (GPCR) that is densely expressed within dopaminergic neurons and circuits. GPCRs are tightly regulated by a variety of intracellular signaling molecules, including Regulator of G Protein Signaling (RGS) proteins. Canonically, RGS proteins act as GTPase accelerating proteins (GAPs) on GTP-bound Ga subunits following GPCR activation, thereby hastening the rate at which GPCR-mediated G protein signaling is terminated. However, some RGS proteins exhibit more complex mechanisms of action on …

Immunomagnetic Beads Coupled With Anti-Ph-20 Antibodies To Isolate Sperm From Dna Mixtures For Sexual Assault Kit Application, Kayla Becks

Graduate Theses, Dissertations, and Problem Reports (ETD)

Sexual Assault Kits (SAKs) account for a large portion of DNA mixture samples that are processed within a forensic laboratory, most commonly in the form of vaginal swabs. A typical vaginal swab from a SAK contains the female victim's epithelial cells and varying concentrations of sperm cells left by the perpetrator. The key to analyzing these types of samples is to separate the mixture into male and female fractions. As such, there is a demand to develop novel techniques that have the capability of recovering as much of the male fraction as possible, specifically in cases where there is a …