Biochemistry, Biophysics, and Structural Biology Commons^™

An Investigation Of Protein Arginine Methyltransferase 1 Substrate Recognition Through Ribonucleoprotein Modifications, Isabella Beverly Lonardo

Undergraduate Honors Capstone Projects

Just as humans need to communicate with each other, our cells have a series of processes that allows them to communicate from one to another. Protein Arginine Methyltransferase I is an enzyme that helps mediate cellular communication by selectively binding to target proteins and performing chemistry that has broad-reaching downstream effects. However, the mechanisms that underly PRMT1 selectivity have yet to be elucidated. This study uses alterations to a series of known targets, called ribonucleoproteins, to identify some of the characteristics that govern recognition by the PRMT1 enzyme. I have made significant progress in the creation, expression, and purification of …

Genetics Of Lrg1 And Its Association With The Cardiometabolic Risk Traits In Mexican American Children And Adolescents, Aaron Rene Reyna

Theses and Dissertations

Leucine-rich alpha-2-glycoprotein 1 (LRG1) has become a major topic of research due to an increasing number of studies demonstrating its unique role in the pathogenesis of multiple human diseases including diabetes and cardiovascular disease. LRG1 has recently been identified as an adipokine, a signaling molecule secreted by adipose tissue. Furthermore, it has been correlated with intercellular signaling that promotes diet-induced hepatosteatosis and insulin resistance in mice. In this study, LRG1 serum concentrations were measured in a family-based cohort of 571 non-diabetic Mexican American children and adolescents aged 6-17 years old (mean age = 11.76 years and females = 48.2%) to …

Effects Of Nitrogen Or Phosphorus Depletion On Lipid Production In Bracteacoccus Sp. And Tetrastrum Sp., Cassandra J. Po

Theses and Dissertations

As global energy demands have risen in the past decades, various species of microalgae have been chosen and used as sources of lipids for third-generation biofuel. I selected two species of green microalgae, Bracteacoccus sp. and Tetrastrum sp., from isolates from South Texas to determine their potential as sources of biofuel. For comparison, Chlamydomonas reinhardtii was used as a model organism. To evaluate lipid productivity under different nutrient regimens, the microalgae were grown in nutrient-replete growth media until the population had reached the end of the logarithmic phase (roughly 14 days), and then transferred into growth media depleted of nitrogen …

Development And Application Of Environmental Dna (Edna) Techniques To Detect Rough-Footed And Yellow Mud Turtles: A Seasonal Field Study To Detect Turtle Edna, Sakib Tahmid Rishan

Theses and Dissertations

The application of environmental DNA (eDNA) has improved the ability of humans to detect the existence and geographical distribution of aquatic organisms. The present study highlights the development and application of eDNA techniques to detect rough-footed mud turtles and yellow mud turtles in different areas in Texas. Both mud turtle species were successfully detected in lakes/resacas using direct eDNA sampling techniques. Moreover, passive eDNA sampling techniques were conducted using different membrane filters to detect mud turtle species. The findings indicated that only the mixed cellulose esters (MCE) filter membranes effectively identified eDNA of yellow mud turtles in the fall, but …

Role Of Interferon-Induced Protein With Tetratricopeptide Repeats 3 (Ifit3) In Hiv-1 Associated Neurodegenerative Disorder And Alzheimer’S Disease, Ranjit Kumar Das

Theses and Dissertations

The advent of cART has revolutionized the management of HIV-1 infection and saved the lives of millions of people worldwide. Yet, HAND continues to be clinically relevant with aging people with HIV-1 (PWH). The underlying mechanism of HAND remains poorly understood. In this regard. Signal transducer and activator of transcription 1(STAT1) and IFIT3 genes have been shown to induce neuroinflammation and neuronal cell death. However, the role of IFIT3 in HAND pathology has not been well established. Therefore, the present work investigated the significant role of IFIT3 gene in development of HAND. In the current study, …

Utilizing Nanodiamond For Antiretroviral Drug Delivery To Hiv Infected Microglial Cells, Ebenezer Owusu

Theses and Dissertations

Combination antiretroviral therapy (cART) is widely used to treat HIV infection, but it fails to eradicate viral reservoirs, such as microglia in the brain, leading to HIV-associated neurological disorders (HAND). Advances in nanomedicine have enabled the development of targeted drug delivery systems, and nanodiamonds (NDs) and fluorescent nanodiamonds (FNDs) have emerged as a potential solution due to their biocompatibility, non-toxic nature, and efficiency as drug carriers. Our study builds on previous findings that NDs can cross the blood-brain barrier (BBB), aiming to utilize NDs and FNDs for their added traceability. We propose a multifunctional nanodrug platform that leverages NDs and …

The Expression Of Recombinant Cas9 Proteins In E. Coli Bl21(De3) Strains, Morgan Tyler Reese

Graduate Theses and Dissertations

Abstract Extensively used for genome editing, CRISPR-Cas9 is a relatively new tool that has made a major impact in the genomic engineering landscape. Due to this, the production of CRISPR proteins that are not commercially available can limit research applications. Here, we present methods for optimizing laboratory scale production of CRISPR proteins including Cas9. The method uses IPTG to induce gene expression of the specified Cas9 and nickel column purification to isolate proteins. CRISPR proteins are validated for purity by SDS-PAGE gel, western blot, and for activity by in vitro digestion. We thoroughly validate and compare the effectiveness of BL21(DE3) …

Exploring The Effects Of N,N-Dimethylbiguanide And Ω-3 Pufas On Muscle Stem Cell Differentiation, Enayah Alhshim

Graduate Theses and Dissertations

The impacts of omega-3 (ω-3) fatty acid and metformin hydrochloride (Met-HCl) on muscle stem cells differentiation, mimicking the fetal muscle development under gestational diabetes mellitus (GDM), were investigated. ω-3 fatty acids have been known for their ability to regulate adipogenic transcription factors such as Sterol Regulatory Element-Binding Protien (SREBPs) and Peroxisome Proliferator-Activated Receptors (PPARs). This regulation resulted in gene expression modification for lipid metabolism and adipocyte differentiation, thus preventing excessive fat accumulation. Additionally, emerging evidence suggested that ω-3 fatty acids may also exert direct effects on myogenic processes that influence muscle stem cells proliferation, differentiation, and maturation. Meanwhile, Met-HCl, an …

Investigation Of How A Kinesin-1 Mutation Affects The Synaptic Localization Of The Periactive Zone Protein Apt-4, Ian Harden

Honors College

Dysfunctional or loss of synaptic transmission is a major contributing factor to many neurological and neurodevelopmental diseases, such as Alzheimer’s disease. To develop novel approaches to treating these diseases, understanding the structures and proteins related to synaptic development and function is essential. Within the synapse is the presynaptic site, containing the active and periactive zones, at which synaptic vesicles containing neurotransmitters are released. The active zone’s primary role is mediating neurotransmitter release via exocytosis, and the periactive zone sorts and recycles synaptic vesicles via endocytosis. However, how periactive zone proteins localize to the presynapse and how they interact with the …

Ecological, Molecular, And Electron Microscopical Investigations Of The Distribution Of Shell-Boring Polychaetes Along The Coast Of Maine, Emma Tomasetti

Honors College

Blister worms are polychaete worms that live in various calcareous materials such as dead shells, red algae, limestone, and cinder blocks, along with cultured shellfish, such as oysters or sea scallops. These worms can be considered a pest due to their nature to burrow into their host’s substrate and create blisters that then fill with mud and detritus. In chapter 1, I worked closely with an oyster farm looking at the reproductive status of one species of blister worm, P. websteri, and the efficacy of cold storage as a method of ridding oysters of P. websteri. For chapter 2, I …

A Combination Of Generated Hydrogen Sulfide And Nitric Oxide Activity Has A Potentiated Protectant Effect Against Cisplatin Induced Nephrotoxicity, Faria Khurshid, Javeid Iqbal, Fiaz Ud Din Ahmad, Arslan Hussain Lodhi, Abdul Malik, Suhail Akhtar, Azmat Ali Khan, Marvi Imam Bux, Mohammed Younis

Biomedical Sciences Faculty Publications

Aim: Hydrogen sulfide and nitricoxide possess cytoprotective activity and in vivo, they are generated from exogenous sodium hydrosulfide and L-arginine respectively. Cisplatin is a major chemotherapeutic agent used to treat cancer and has a high incidence of nephrotoxicity as a side effect. The study aim was to explore the effects of NaHS and L-arginine or their combination on cisplatin induced nephrotoxicity in rats. Methods: Wistar Kyoto rats were given a single intraperitoneal dose of cisplatin (5 mg/kg) followed either by NaHS (56 μmol/kg, i. p.), L-arginine (1.25 g/L in drinking water) or their combination daily for 28-days. Post-mortem plasma, urine …

The Purification And Thermal Stability Of The Peroxidase Enzyme In Cucurbita Moschata, Garen Hamner

Senior Honors Theses

Peroxidases are enzymes that catalyze the reduction of hydrogen peroxide to water while oxidizing organic substrates and are valuable in spheres like industrial and medical applications and histochemistry. Limitations still exist in the use of the well-studied horseradish peroxidase for certain activities due to limitations like poor thermal stability, thus the search for novel peroxidases that can overcome these limitations is an active area of research. Butternut squash peroxidase (Cucurbita moschata) (BSP) shows promise due to significant activity being found in the skin and apparent enhanced thermal stability, but an efficient purification scheme for it is lacking, as well as …

Using Edna To Assess Impacts Of Oyster Restoration On Ecosystem Biodiversity At A Heavily Impacted Coastal Lagoon, Dominique Di Domenico, Emily Bonacchi, Elizabeth Suter

Molloy Multidisciplinary Undergraduate Research Conference

The Great South Bay (GSB), a bar-built, temperate, coastal lagoon on the south shore of Long Island, New York, USA, is a highly modified estuary due to heavy urbanization and suburbanization in the last century. GSB historically provided up to 50% of the nation’s hard clam (Mercenaria mercenaria) harvest, and is the legal home of the “Blue Point” oyster (eastern oyster; Crassostrea virginica). However, due to overexploitation as well as persistent stress from ongoing nitrogen pollution and harmful algal blooms, shellfish populations are fractions of their original abundances. Restoration efforts in GSB, driven both by community efforts …

Investigating The Role Of Tryptophan In The Native Photo-Crosslinking Of A Cys-Containing Transmembrane Helix To A Membrane Lipid Double Bond, Michael Jay Telehany

Undergraduate Honors Theses

Our research group has previously discovered that a Cys residue within a transmembrane (TM) helix can directly photo-crosslink to a membrane lipid via an oxythiolation addition reaction to the double bond within the lipid, with a reaction yield of 10-20%. The pH-low Insertion Peptide (pHLIP) serves as our model TM helix, while the POPC lipid bilayer is our model membrane. We had long believed that the Trp residues within the sequence of pHLIP served as the key chromophore for this reaction, and thus they were essential for the formation of the TM helix-lipid adduct. To test this hypothesis, I synthesized …

Trna Anticodon Cleavage By Target-Activated Crispr-Cas13a Effector, Ishita Jain, Matvey Kolesnik, Konstantin Kuznedelov, Leonid Minakhin, Natalia Morozova, Anna Shiriaeva, Alexandr Kirillov, Sofia Medvedeva, Alexei Livenskyi, Laura Kazieva, Kira S Makarova, Eugene V Koonin, Sergei Borukhov, Konstantin Severinov, Ekaterina Semenova

Rowan-Virtua School of Osteopathic Medicine Departmental Research

Type VI CRISPR-Cas systems are among the few CRISPR varieties that target exclusively RNA. The CRISPR RNA–guided, sequence-specific binding of target RNAs, such as phage transcripts, activates the type VI effector, Cas13. Once activated, Cas13 causes collateral RNA cleavage, which induces bacterial cell dormancy, thus protecting the host population from the phage spread. We show here that the principal form of collateral RNA degradation elicited by Leptotrichia shahii Cas13a expressed in Escherichia coli cells is the cleavage of anticodons in a subset of transfer RNAs (tRNAs) with uridine-rich anticodons. This tRNA cleavage is accompanied by inhibition of protein synthesis, thus …

Modeling And Synthesis Of 3-(2,6-Disubstituted 5-Pyrimidyl) Propionic Acids As Inhibitors Of Low Molecular Weight Protein Tyrosine Phosphatase (Lmw-Ptp), Kennedy Bueno, Madisen Carter

Celebrating Scholarship and Creativity Day (2018-)

Low molecular weight protein tyrosine phosphatase (LMW-PTP) is an enzyme and a known signal pathway for growth factors and cellular transformation in eukaryotic cells. Our research aims to synthesize 12 new potential inhibitors of LMW-PTP and analyze their inhibitory activity and binding affinity. The inhibition of both isoforms of LMW-PTP has been the primary focus of research due to their potential role in breast, colon, and other cancers as well as type II diabetes. One known inhibitor, pyridoxal 5’- phosphate (PLP), is essential for various enzymatic reactions within the body, such as the synthesis of neurotransmitters, and is therefore impractical …

Improving The Drug Antipyrine: Synthesis Of N2-Aryl Analogs Through Oxidation Of Dihydroantipyrines, Eli Mans

Student Academic Conference

Idiopathic Pulmonary Fibrosis (IPF) is a progressive and fatal lung disease that affects thousands of people worldwide. Current drugs are used in clinical settings to treat IPF but these drugs have low efficacy and a high monetary costs. The molecule antipyrine has been identified by colleagues at Mayo Clinic as an early-stage drug candidate for treatment for IPF. The Jasperse group is synthesizing analogs of antipyrine to create a drug library for improved IPF treatment. A novel three step process via a silyl enol ether intermediate has been developed to convert a C-C bond to a C=C bond between C4-C5 …

Predictive And Prognostic Biomarkers And Tumor Antigens For Targeted Therapy In Urothelial Carcinoma, Aditya Eturi, Amman Bhasin, Kevin Zarrabi, William Tester

Department of Medical Oncology Faculty Papers

Urothelial carcinoma (UC) is the fourth most prevalent cancer amongst males worldwide. While patients with non-muscle-invasive disease have a favorable prognosis, 25% of UC patients present with locally advanced disease which is associated with a 10-15% 5-year survival rate and poor overall prognosis. Muscle-invasive bladder cancer (MIBC) is associated with about 50% 5 year survival when treated by radical cystectomy or trimodality therapy; stage IV disease is associated with 10-15% 5 year survival. Current therapeutic modalities for MIBC include neoadjuvant chemotherapy, surgery and/or chemoradiation, although patients with relapsed or refractory disease have a poor prognosis. However, the rapid success of …

Isolation And Analysis Of R130d Malate Dehydrogenase (Mdh), Hayley Brunette, Abby Doyle, Ilsa Hoaglund, Danielle Bolland

Undergraduate Research Symposium 2024

Malate Dehydrogenase (MDH) is the enzyme that catalyzes the ninth step of the citric acid cycle, a regenerative step in the metabolism of glucose in which malate is converted to oxaloacetate. Conformational changes in the active loop region of MDH along with the coenzyme NAD+ drive catalysis. Previous studies suggested that mutating positively charged amino acids to negatively charged amino acids at residue 130 decreased the enzymatic activity of MDH. We hypothesized that if we change arginine 130 (positively charged amino acid) to aspartate (negatively charged amino acid), then the enzymatic activity of MDH would significantly decrease. In e. Coli …

Upregulation Of The Predominant Cystic Fibrosis Causing Mutation Df508-Cftr By Triazole Compounds In Epithelial Cells, Maggie Taylor

Undergraduate Research Conference

Cystic fibrosis is a common genetic disease that is caused by a mutation in the plasma membrane protein CFTR, which stands for Cystic Fibrosis Transmembrane-conductance Regulator. When this membrane protein is mutated, it impairs its chloride ion channel function, blocking the movement of chloride ions that travel in and out of the cell. Previous studies have demonstrated that the most prevalent CFTR mutation, ∆F508-CFTR, can be partially reversed using small molecules (Heda and Marino, BBRC, 271:659-664, 2000). In this study, I have investigated the effects of several triazole compounds known to bind and transport chloride ions in cultured cells, on …

Hgs-3 The Influence Of A Tandem Cycling Program In The Community On Physical And Functional Health, Therapeutic Bonds, And Quality Of Life For Individuals And Care Partners Coping With Parkinson’S Disease, Leila Djerdjour, Jennifer L. Trilk

SC Upstate Research Symposium

Purpose Statement: Several studies have shown that aerobic exercise can have a positive impact on alleviating symptoms experienced by individuals with Parkinson's disease (PD). Despite this evidence, the potential benefits of exercise for both PD patients and their care partners (PD dyad) remain unexplored. This research project investigates the effectiveness, therapeutic collaborations, and physical outcomes of a virtual reality (VR) tandem cycling program specifically designed for PD dyads.

Methods: Following approval from the Prisma Health Institutional Review Board, individuals with PD were identified and screened by clinical neurologists. The pre-testing measures for PD dyads (N=9) included emotional and cognitive status …

Multivalency Of The Mdmx Acidic Domain Slims Use Different Binding Modes To Inhibit Mdmx And P53, Malissa Fenton

USF Tampa Graduate Theses and Dissertations

Regulation of protein binding through autoinhibition commonly occurs via interactions involving intrinsically disordered regions (IDRs). These intramolecular interactions can directly or allosterically inhibit intermolecular protein or DNA binding, regulate enzymatic activity, and control the assembly of large macromolecular complexes. Autoinhibitory interactions mediated by protein disorder are inherently transient, making their identification and characterization challenging. In this work, we explore the structural and functional diversity of disorder-mediated autoinhibition for a variety of biological mechanisms, with a focus on the role of multivalency and effective concentration. We also discuss the evolution of disordered motifs that participate in autoinhibition using examples where sequence …

Lysine 473 Regulates The Activity And Trafficking Of The Cystine/Glutamate Transporter, System XC-, Anna Koppin

23rd Annual A. Paul and Carol C. Schaap Celebration of Undergraduate Research and Creative Activity (2024)

System x_c^- is a membrane transport system that plays a critical role in mitigating oxidative stress. Past work in our lab has shown that System x_c^- localizes to the plasma membrane allowing for increased activity to support production of antioxidants during oxidative stress. In this study, we sought to determine if post-translational modification (PTM) of the transporter regulates its trafficking. A C-terminal 3KR mutant (K422,472,473R) exhibited decreased membrane localization and activity, suggesting that PTM at one of these sites increases activity. Further, we observed that K473R exhibits a 7 kD decrease in the molecular weight, indicating …

Lc-Ms/Ms For Proteomic Analysis Of Post-Translational Modifications On Xct, Connor Bricco

23rd Annual A. Paul and Carol C. Schaap Celebration of Undergraduate Research and Creative Activity (2024)

Membrane protein xCT and its heavy chain component 4F2HC make up the xc- transport system. 4F2HC may be necessary for membrane localization of the heterodimer and xCT is responsible for transport activity. Under basal conditions, xCT resides in endosomes, but upon oxidative insult, xCT moves to the membrane and functions to reduce oxidative stress. We hypothesize the movement of xCT to the membrane is directed by changes in posttranslational modifications (PTMs) such as phosphorylation, ubiquitination and glycosylation. The overall goal of this project is to use mass spectrometry to detect the PTMs of xCT isolated from cells grown under basal …

Ubiquitination Of Xct: Impacts On The Protein's Stability, Turnover Rate, And Localization, Sofia Rosenberger

23rd Annual A. Paul and Carol C. Schaap Celebration of Undergraduate Research and Creative Activity (2024)

System x_c^- imports cystine and exports glutamate. Its presence on the plasma membrane has been shown to increase directly with oxidative insults. Ubiquitin, a small protein, is directly involved in the trafficking and degradation of numerous proteins within cells and has been shown to bind to System x_c^-. Moreover, upon oxidative insult, protein ubiquitination increases. However, it is not understood how ubiquitination of the transporter impacts its activity. Therefore, the objective of this project is to directly assess how ubiquitination affects the protein’s stability, turnover rate, and localization in the context of oxidative stress. We …

Using Artificial (Ai) To Predict A Structure Of Protein Complex, Yiqing Zang

SACAD: John Heinrichs Scholarly and Creative Activity Days

Proteins play pivotal roles in essential life processes and elucidating their three-dimensional (3D) structures is crucial for understanding their functions. AlphaFold2, an advanced artificial intelligence-based method developed by Google DeepMind, has emerged as a promising tool for predicting protein structures. In this study, we evaluated the predictive capabilities of AlphaFold2. Our findings highlight AlphaFold2's efficacy in providing valuable insights into protein structure prediction, albeit with certain limitations. While AlphaFold2 represents a significant advancement in the field, its utility is best realized when integrated with complementary experimental approaches. Consequently, combining the strengths of AlphaFold2 with experimental validation remains essential for achieving …

Discovery Of A Small-Molecule Inhibitor That Traps Polθ On Dna And Synergizes With Parp Inhibitors, William Fried, Mrityunjay Tyagi, Leonid Minakhin, Gurushankar Chandramouly, Taylor Tredinnick, Mercy Ramanjulu, William Auerbacher, Marissa L Calbert, Timur Rusanov, Trung Hoang, Nikita Borisonnik, Robert Betsch, John Krais, Yifan Wang, Umeshkumar Vekariya, John Gordon, George Morton, Tatiana Kent, Tomasz Skorski, Neil Johnson, Wayne Childers, Xiaojiang Chen, Richard Pomerantz

Department of Biochemistry and Molecular Biology Faculty Papers

The DNA damage response (DDR) protein DNA Polymerase θ (Polθ) is synthetic lethal with homologous recombination (HR) factors and is therefore a promising drug target in BRCA1/2 mutant cancers. We discover an allosteric Polθ inhibitor (Polθi) class with 4-6 nM IC₅₀ that selectively kills HR-deficient cells and acts synergistically with PARP inhibitors (PARPi) in multiple genetic backgrounds. X-ray crystallography and biochemistry reveal that Polθi selectively inhibits Polθ polymerase (Polθ-pol) in the closed conformation on B-form DNA/DNA via an induced fit mechanism. In contrast, Polθi fails to inhibit Polθ-pol catalytic activity on A-form DNA/RNA in which the enzyme binds in …

The Impact Of Mutations In The Arabidopsis Apetela (Ap3) Gene, Hazel R. Frans, Tara Phelps-Durr

SACAD: John Heinrichs Scholarly and Creative Activity Days

The purpose of this research is to understand the molecular functioning of the Arabidopsis thaliana Apetela (Ap3) gene. We created mutations in two sites of the gene, AP3-3 and AP3-5. These are predicted to change AP3 protein structure, which may result in a mutated flower. Analyzing the effects of new mutations allows an understanding of protein formation both in plants and humans.

Social Media Does Not Elicit A Physiological Stress Response As Measured By Heart Rate And Salivary Cortisol Over 20-Minute Sessions Of Cell Phone Use, Suzanne Oppenheimer, Laura Bond, Charity Smith

Biomolecular Research Institute Publications and Presentations

The pervasive use of social media has raised concerns about its potential detrimental effects on physical and mental health. Others have demonstrated a relationship between social media use and anxiety, depression, and psychosocial stress. In light of these studies, we examined physiological indicators of stress (heart rate to measure autonomic nervous system activation and cortisol to assess activity of the hypothalamic-pituitary-adrenal axis) associated with social media use and investigated possible moderating influences of sex, age, and psychological parameters. We collected physiological data from 59 subjects ranging in age from 13 to 55 across two cell phone treatments: social media use …

Parp2 Promotes Break Induced Replication-Mediated Telomere Fragility In Response To Replication Stress, Daniela Muoio, Natalie Laspata, Rachel L Dannenberg, Caroline Curry, Simone Darkoa-Larbi, Mark Hedglin, Shikhar Uttam, Elise Fouquerel

Department of Biochemistry and Molecular Biology Faculty Papers

PARP2 is a DNA-dependent ADP-ribosyl transferase (ARTs) enzyme with Poly(ADP-ribosyl)ation activity that is triggered by DNA breaks. It plays a role in the Base Excision Repair pathway, where it has overlapping functions with PARP1. However, additional roles for PARP2 have emerged in the response of cells to replication stress. In this study, we demonstrate that PARP2 promotes replication stress-induced telomere fragility and prevents telomere loss following chronic induction of oxidative DNA lesions and BLM helicase depletion. Telomere fragility results from the activity of the break-induced replication pathway (BIR). During this process, PARP2 promotes DNA end resection, strand invasion and BIR-dependent …