Biochemistry, Biophysics, and Structural Biology Commons^™

Molecular Mass Of L-Arginine-Based Potentially Carcinogenic Heterocyclic Amines, Irene S. Hwang

Honors Theses

Investigation of cancer-inducing molecules in cooked foods has led to the discovery of mutagenic heterocyclic amines (HCAs) in meat. The amino acids creatin(in)e and L-phenylalanine form the precursors for these HCAs. Recent research here at Andrews University have found similar mutagenic HCAs produced from plant-based proteins when L-arginine, rather than creatin(in)e, is substituted in high temperature reactions (simulated cooking) with L-phenylalanine. Our research focused on developing HPLC methods to isolate individual candidates that are then screened for mutagenicity via the Ames test. Mass spectroscopy methods were also developed to identfy the molecular structure of these lead mutagenic L-arginine-based HCAs.

Fluorescence Analysis Of Pamam Dendrimers, Ansel Nam

Honors Theses

Despite their lack of conjugation, polyamidoamine (PAMAM) dendrimers are intrinsically fluorescent, and this fluorescent is somewhat manipulable through altering the dendrimer's solvent. This level of fluorescence is dependent on a number of the dendrimer's qualities: the pH of the solvent, the mass percent concentration of the dendrimer in solvent, and the generation size of the dendrimer. In past studies, fluorescent intensity increased as generation size increased and pH decreased. Thus, this research project will attempt to replicate those results as well as further analyze the efficiency of fluorescence of PAMAM dendrimer in comparison to fluorescein.

Mutagenic And Spectroscopic Investigation Of Ph Dependent Cooa Dna Binding, Brian R. Weaver

Chemistry Honors Papers

The carbon monoxide (CO) sensing heme protein, CooA, is a transcription factor which exists in several bacteria that utilize CO as an energy source. CooA positively regulates the expression of coo genes in the presence of CO such that the corresponding proteins may metabolize CO. The present studies have yielded the unexpected result that Fe(III) CooA binds DNA tightly at pH < 7, deviating from all previously reported work which indicate that CooA DNA binding is initiated only when the exogenous CO effector reacts with the Fe(II) CooA heme. This observation suggests that the disruption of one or more salt bridges upon effector binding may be a critical feature of the normal CooA activation mechanism. To test this possibility, several protein variants that eliminated a selected salt bridge for the CooA homolog from Rhodospirillum rubrum were prepared via site-directed mutagenesis. Samples of these variant proteins, which were overexpressed in Escherichia coli, were then characterized by spectroscopic methods and functional assays to investigate the impact these mutations had on CooA heme coordination …

International Collaboration And New Technology Reveal Rare Genes That Determine Height, Nicolle Dauphinee

Thinking Matters Symposium Archive

Height is a phenotype, an observable trait of the genes that we inherit from our parents. It's a polygenic trait, meaning it is controlled by 2 or more genes. Because it is so easy to measure, it is an ideal characteristic to help us understand other polygenic traits. So far, genome-wide association studies (GWAS) have identified 700 common variants that determine a person's height. New discoveries have been made by the international group called Genetic Investigation of Anthropometric Traits (GIANT), revealing 83 genetic mutations that have additional effects on a person's height.¹

The Effects Of Carrier Ligands On Cisplatin Binding To Cysteine And Methionine, Adam C.R Smith

Masters Theses & Specialist Projects

We have reacted several derivatives of the anticancer drug cisplatin with N-acetyl-Lcysteine (N-AcCys) and N-acetyl-L-methionine (N-AcMet), which are two of the primary amino acid targets of platinum. NMR spectroscopy was used to monitor the reactions and determine the effect the different ligands would have on the platinum reactivity. Several of the platinum compounds were tested at pH of 4 and 7, and with platinum:amino acid ratios of 1:1, 2:1 and 1:2. Competition reactions between cysteine and methionine were done to confirm which would react with the platinum compound first. [Pt(dien)(NO3)]+ reacts faster with methionine than with cysteine at both pH …

The Statistical Dynamics Of Nonequilibrium Control, Grant Murray Rotskoff '09

Doctoral Dissertations

Living systems, even at the scale of single molecules, are constantly adapting to changing environmental conditions. The physical response of a nanoscale system to external gradients or changing thermodynamic conditions can be chaotic, nonlinear, and hence difficult to control or predict. Nevertheless, biology has evolved systems that reliably carry out the cell’s vital functions efficiently enough to ensure survival. Moreover, the development of new experimental techniques to monitor and manipulate single biological molecules has provided a natural testbed for theoretical investigations of nonequilibrium dynamics. This work focuses on developing paradigms for both understanding the principles of nonequilibrium dynamics and also …

Analysis Of Hiv-1 Quasispecies Sequences Generated By High Throughput Sequencing (Hts) Using Hive, Naila Gulzar, Bhavna Hora, Konstantinos Karagiannis, Krista Smith, Feng Gao, Raja Mazumder

GW Research Days 2016 - 2020

The high level of genetic variability of Human Immunodeficiency Virus type 1 (HIV-1) is caused by the low fidelity of its replication machinery. This leads to evolution of swarm-like viral populations often described as quasispecies. High throughput sequencing (HTS) technology provides higher resolution over Sanger sequencing, enabling detection of low frequency variant genomes. However, quasispecies analysis is still a challenge due to the systematic noise, introduced by HTS technology. This leads to the increase in type I errors (also known as false positives) and the underlying genetic diversity, which can lead to mathematically insolvable type II errors (also known as …

Development Of An In Vivo Assay For Acetyl-Coa Carboxylase Using 1,3,6,8- Tetrahydroxynapthalene Synthase, Sarah Bond

Honors Capstones

No abstract provided.

Synthesis, Kinetic And Catalytic Studies Of Manganese Complexes With Corrole And Porphyrin Ligands, Haleh Jeddi

Masters Theses & Specialist Projects

High-valent transition metal-oxo intermediates play a significant role in the catalytic cycle of the ubiquitous cytochrome P450 enzymes and in biomimetic catalytic systems. In this work, manganese(III) porphyrin and corrole systems (2) were synthesized and characterized by UV-vis absorbance and 1H-NMR, matching literaturereported spectroscopic data. Manganese(V)-oxo corroles (3) and a manganese(IV)-oxo porphyrin (4) were successfully generated by chemical oxidation using mchloroperoxybenzoic acid (m-CPBA), and their oxidation reactions with organic reductants were comparatively investigated. Results from single-turnover kinetic studies indicate that in the tris(pentafluorophenyl)corrole system (3a), the active oxidizing intermediate differs in different solvents. The active oxidizing intermediate in acetonitrile is …

Effects Of Binding Affinity Between Bovine Serum Albumin And Platinum Drugs, Nathan Puckett

Masters Theses & Specialist Projects

Platinum complex drugs such as cisplatin have been used as highly successful chemotherapy drugs since the 1970s. We are interested in how the ligands attached to cisplatin analogs influences their reactivity with biologically relevant targets along with time and amount. For this study, reactions were conducted to determine the reactivity between different platinum compounds and the protein bovine serum albumin. Various platinum compounds with different ligands were reacted in varying amounts with albumin in ammonium acetate buffer for either 1 hour, 4 hours, or 24 hours. Each reaction was quenched after the designated reaction time by dialysis and the platinum …

Correlated Mutation In The Evolution Of Catalysis In Uracil Dna Glycosylase Superfamily, Bo Xia, Yingling Liu, Jose Guevara, Jing Li, Celeste Jilich, Ye Yang, Liangjiang Wang, Brian N. Dominy, Weiguo Cao

Publications

Enzymes in Uracil DNA glycosylase (UDG) superfamily are essential for the removal of uracil. Family 4 UDGa is a robust uracil DNA glycosylase that only acts on double-stranded and single-stranded uracil-containing DNA. Based on mutational, kinetic and modeling analyses, a catalytic mechanism involving leaving group stabilization by H155 in motif 2 and water coordination by N89 in motif 3 is proposed. Mutual Information analysis identifies a complexed correlated mutation network including a strong correlation in the EG doublet in motif 1 of family 4 UDGa and in the QD doublet in motif 1 of family 1 UNG. Conversion of EG …

Computing Spatiotemporal Heat Maps Of Lipid Electropore Formation: A Statistical Approach, Willy Wriggers, Frederica Castellani, Julio A. Kovacs, P. Thomas Vernier

Mechanical & Aerospace Engineering Faculty Publications

We extend the multiscale spatiotemporal heat map strategies originally developed for interpreting molecular dynamics simulations of well-structured proteins to liquids such as lipid bilayers and solvents. Our analysis informs the experimental and theoretical investigation of electroporation, that is, the externally imposed breaching of the cell membrane under the influence of an electric field of sufficient magnitude. To understand the nanoscale architecture of electroporation, we transform time domain data of the coarse-grained interaction networks of lipids and solvents into spatial heat maps of the most relevant constituent molecules. The application takes advantage of our earlier graph-based activity functions by accounting for …

A Balanced Approach To Adaptive Probability Density Estimation, Julio Kovacs, Cailee Helmick, Willy Wriggers

Mechanical & Aerospace Engineering Faculty Publications

Our development of a Fast (Mutual) Information Matching (FIM) of molecular dynamics time series data led us to the general problem of how to accurately estimate the probability density function of a random variable, especially in cases of very uneven samples. Here, we propose a novel Balanced Adaptive Density Estimation (BADE) method that effectively optimizes the amount of smoothing at each point. To do this, BADE relies on an efficient nearest-neighbor search which results in good scaling for large data sizes. Our tests on simulated data show that BADE exhibits equal or better accuracy than existing methods, and visual tests …

Quantifying Cyanide Inhibition Of Nitrification And Developing Cost-Effective Treatment Processes, Germano M. Salazar-Benites

Civil & Environmental Engineering Theses & Dissertations

All wastewater treatment plants that operate multiple hearth furnaces (MHF) and are required to nitrify must manage the inhibitory effects of free cyanide (HCN, CN-) in the scrubber return flows due to inhibitory impacts on nitrifying bacteria.

HRSD Boat Harbor Treatment Plant (BHTP) a 25 MGD facility consisting of primary and secondary treatment, employs an anoxic selector process for nitrification and partial denitrification and operates a MHF. There is a desire to improve TN removal performance at BHTP due to an annual mass-based bubble permit limit on a combined discharge from seven HRSD plants, and there are no discharge limitations …

Natural Killer (Nk) Cells And Their Involvement In Different Types Of Cancer. Current Status Of Clinical Research, Isadora Zaharescu, Adina D. Moldovan, Cristiana Tanase

Journal of Mind and Medical Sciences

Natural killer cells are the main agents of innate immunity. Since 1970, various studies have repeatedly confirmed their involvement in decreasing local tumor growth and also decreasing the risk of metastasis, due to their cytotoxic effects and also through the release of immunostimulatory cytokines such as IFN-gamma. In the 1990s, several studies demonstrated the existence of certain inhibiting and stimulating receptors of these cells, leading to the concept of “induced self”, thus explaining why tumors with MHC-1 are destroyed and autologous cells without it are saved out. Recognition and destruction of tumor cells by the NK cells are the result …

Reactivity Of Ketyl And Acetyl Radicals From Direct Solar Actinic Photolysis Of Aqueous Pyruvic Acid, Alexis J. Eugene, Marcelo I. Guzman

Chemistry Faculty Publications

The variable composition of secondary organic aerosols (SOA) contributes to the large uncertainty for predicting radiative forcing. A better understanding of the reaction mechanisms leading to aerosol formation such as for the photochemical reaction of aqueous pyruvic acid (PA) at λ ≥ 305 nm can contribute to constrain these uncertainties. Herein, the photochemistry of aqueous PA (5-300 mM) continuously sparged with air is re-examined in the laboratory under comparable irradiance at 38° N at noon on a summer day. Several analytical methods are employed to monitor the time series of the reaction, including (1) the derivatization of carbonyl (C═O) functional …

Exploring The Use Of Free Bioinformatics Modules In An Introductory Biochemistry Course, Charlsey Dodgen, Uzezi Uwerosuo, Chulhee Kang, Cathy Lee

Georgia Journal of Science

Although bioinformatics, the use of computational science to study biology, has become imperative in many areas of the biological sciences and related career paths, introductory biochemistry courses may disregard practical knowledge on bioinformatics. For this reason, we merged a hands-on activity module into an undergraduate biochemistry course in two ways. First, we incorporated bioinformatics modules for building phylogenetic trees by aligning the active sites of 10 chosen related α-amylase enzymes using freely available data. Secondly, we chose three of those 10 α-amylase enzymes to compare the 3D structure of their active sites. This module gives the students an opportunity to …

Another Look At Pyrroloiminoquinone Alkaloids-Perspectives On Their Therapeutic Potential From Known Structures And Semisynthetic Analogues., Sheng Lin, Erin P. Mccauley, Nicholas Lorig-Roach, Karen Tenney, Cassandra N. Naphen, Ai-Mei Yang, Tyler A. Johnson, Thalia Hernadez, Ramandeep Rattan, Frederick A. Valeriote, Phillip Crews

Natural Sciences and Mathematics | Faculty Scholarship

This study began with the goal of identifying constituents from Zyzzya fuliginosa extracts that showed selectivity in our primary cytotoxicity screen against the PANC-1 tumor cell line. During the course of this project, which focused on six Z. fuliginosa samples collected from various regions of the Indo-Pacific, known compounds were obtained consisting of nine makaluvamine and three damirone analogues. Four new acetylated derivatives were also prepared. High-accuracy electrospray ionization mass spectrometry (HAESI-MS) m/z ions produced through MS² runs were obtained and interpreted to provide a rapid way for dereplicating isomers containing a pyrrolo[4,3,2-de]quinoline core. In vitro human pancreas/duct epithelioid …

Gene 33/Mig6 Regulates Apoptosis And The Dna Damage Response Through Independent Mechanisms, Cen Li, Soyoung Park, Leonard M. Eisenberg, Hong Zhao, Zbigniew Darzynkiewicz, Dazhong Xu

NYMC Faculty Posters

Gene 33 (Mig6, ERRFI1) is an inducible adaptor/scaffold protein whose expression can be induced by both stress and mitogenic signals. It contains multiple domains for protein-protein interaction and is involved in a broad spectrum of cellular functions. Gene 33 promotes apoptosis in a cell type-dependent manner. A recent study has linked Gene 33 to the DNA damage response (DDR) induced by hexavalent chromium [Cr(VI)]. Here we show that Gene 33 induces apoptosis via both c-Abl/p73 and EGFR/AKT-dependent pathways in lung epithelial and lung carcinoma cells. Ectopic expression of Gene 33 also triggers DDR in an ATM-dependent fashion and through pathways …

Plp And Gaba Trigger Gabr-Mediated Transcription Regulation In Bacillus Subtilis Via External Aldimine Formation, Rui Wu, Ruslan Sanishvili, Boris R. Belitsky, Jose I. Juncosa, Hoang V. Le, Helaina J. S. Lehrer, Michael Farley, Richard B. Silverman, Gregory A. Petsko, Dagmar Ringe, Dali Liu

Chemistry: Faculty Publications and Other Works

The Bacillus subtilis protein regulator of the gabTD operon and its own gene (GabR) is a transcriptional activator that regulates transcription of γ-aminobutyric acid aminotransferase (GABA-AT; GabT) upon interactions with pyridoxal-5′-phosphate (PLP) and GABA, and thereby promotes the biosynthesis of glutamate from GABA. We show here that the external aldimine formed between PLP and GABA is apparently responsible for triggering the GabR-mediated transcription activation. Details of the “active site” in the structure of the GabR effector-binding/oligomerization (Eb/O) domain suggest that binding a monocarboxylic γ-amino acid such as GABA should be preferred over dicarboxylic acid ligands. A reactive GABA analog, ( …

Mass-Spectrometry Based Proteomics Of Age-Related Changes In Murine Microglia, Antwoine Flowers

USF Tampa Graduate Theses and Dissertations

The last century has seen a steady increase in the extension of the average lifespan. This has concomitantly produced higher incidences of age-related chronic degenerative diseases like Alzheimer’s and Parkinson’s diseases. Age is the single greatest risk factor for the development of not just these degenerative conditions but cancer as well. The aged niche undergoes a number of maladaptive changes that allow underlying conditions to present and progress. Exactly which changes, contribute to the progression of which disease is currently an area of intense study. However, these answers often present therapeutic targets for disease prevention. Age is characterized by a …

Hydrogen Peroxide Inhibition And Inactivation Of Bicupin Oxalate Oxidase, John M. Goodwin Vii

Master of Science in Chemical Sciences Theses

Oxalate oxidase is a manganese containing enzyme that catalyzes the oxygen dependent oxidation of oxalate to carbon dioxide and hydrogen peroxide. Oxalate oxidase from Ceriporiopsis subvermispora (CsOxOx) is the first characterized bicupin oxalate oxidase. Potential applications of oxalate oxidase in cancer treatment, paper pulping, and biofuel cells highlight the need to study the effects of hydrogen peroxide on the CsOxOx catalyzed oxidation of oxalate. Membrane inlet mass spectrometry (MIMS) was used to directly measure initial rates of carbon dioxide formation and oxygen consumption in varying initial concentrations of hydrogen peroxide. This work demonstrates that hydrogen peroxide is both a reversible …

Hierarchical Corannulene-Based Materials: Energy Transfer And Solid-State Photophysics, Allison M. Rice, W. Brett Fellows, Ekaterina A. Dolgopolova, Andrew B. Greytak, Aaron K. Vannucci, Mark D. Smith, Stavros G. Karakalos, Jeanette A. Krause, Stanislav M. Avdoshenko, Alexey A. Popov, Natalia B. Shustova Prof. Dr.

Faculty Publications

We report the first example of a donor–acceptor corannulene-containing hybrid material with rapid ligand-to-ligand energy transfer (ET). Additionally, we provide the first time-resolved photoluminescence (PL) data for any corannulene-based compounds in the solid state. Comprehensive analysis of PL data in combination with theoretical calculations of donor–acceptor exciton coupling was employed to estimate ET rate and efficiency in the prepared material. The ligand-to-ligand ET rate calculated using two models is comparable with that observed in fullerene-containing materials, which are generally considered for molecular electronics development. Thus, the presented studies not only demonstrate the possibility of merging the intrinsic properties of π-bowls, …

Development Of Tools For Stable Transfection In The Human Filarial Parasite Brugia Malayi Via The Piggybac Transposon System, Johan Chabanon

USF Tampa Graduate Theses and Dissertations

Brugia malayi is one of three species of nematode known to cause lymphatic filariasis (LF) in humans. LF infects over 120 million people, causing debilitating disease. Various global programs have been launched in the past 20 years to eliminate LF. These programs have greatly scaled up the resources and efforts allocated to halting the transmission and reducing disease burden. Only a few drugs are used to treat LF, and resistance is thus a devastating possibility. Research aimed at identifying new drug targets could therefore prove essential in elimination of LF.

Genetic manipulation of B. malayi has been limited to transient …

A Novel Abi-Domain Protein Controls Virulence Determinant Production In Staphylococcus Aureus, Stephanie Michelle Marroquin

USF Tampa Graduate Theses and Dissertations

A major factor in the success of Staphylococcus aureus as a pathogen is its vast arsenal of virulence determinants and, more importantly, the tight and precisely- timed regulation of these factors. Here we investigate the product of the S. aureus gene, SAUSA300_1984, encoding a putative transmembrane protein. This as yet uncharacterized protein belongs to the Abi (abortive infection) family, which are commonly annotated as CAAX-proteases, and are significantly understudied in prokaryotes. In S. aureus the disruption of SAUSA300_1984 results in a drastic reduction of proteolytic and hemolytic activity, as well as diminished pigmentation. This phenotype appears to be mediated through …

Microbal Strans And Methods Of Making And Using, Nicole Roswitha Buan Murphy, Jennifer Catlett

Department of Biochemistry: Faculty Publications

Microbial strains are provided, as are methods of making and using Such microbial strains.

Characterization Of Lipid Rafts In Human Platelets Using Nuclear Magnetic Resonance: A Pilot Study, Joshua F. Ceñido, Boris Itin, Ruth E. Stark, Yung-Yu Huang, Maria A. Oquendo, J. John Mann, M. Elizabeth Sublette

Publications and Research

Lipid microdomains (‘lipid rafts’) are plasma membrane subregions, enriched in cholesterol and glycosphingolipids, which participate dynamically in cell signaling and molecular trafficking operations. One strategy for the study of the physicochemical properties of lipid rafts in model membrane systems has been the use of nuclear magnetic resonance (NMR), but until now this spectroscopic method has not been considered a clinically relevant tool. We performed a proof-of-concept study to test the feasibility of using NMR to study lipid rafts in human tissues. Platelets were selected as a cost-effective and minimally invasive model system in which lipid rafts have previously been studied …

Discovery And Validation Of Information Theory-Based Transcription Factor And Cofactor Binding Site Motifs., Ruipeng Lu, Eliseos J Mucaki, Peter K Rogan

Biochemistry Publications

Data from ChIP-seq experiments can derive the genome-wide binding specificities of transcription factors (TFs) and other regulatory proteins. We analyzed 765 ENCODE ChIP-seq peak datasets of 207 human TFs with a novel motif discovery pipeline based on recursive, thresholded entropy minimization. This approach, while obviating the need to compensate for skewed nucleotide composition, distinguishes true binding motifs from noise, quantifies the strengths of individual binding sites based on computed affinity and detects adjacent cofactor binding sites that coordinate with the targets of primary, immunoprecipitated TFs. We obtained contiguous and bipartite information theory-based position weight matrices (iPWMs) for 93 sequence-specific TFs, …

Role Of Inflammation In 20-Hete Regulation Of Ischemia-Induced Angiogenesis, Elizabeth Berry, Rachel John, Samantha Tang, Austin M. Guo

NYMC Faculty Posters

Objective: 20-Hydroxyeicosatetraenoic acid (20-HETE), an important bioactive lipid metabolite, has recently been identified to be a novel contributor of angiogenesis secondary to ischemia. Moreover, an inflammatory response is required for the initiation of ischemic angiogenesis, in response to ischemic tissue injury. The goal of this study is to investigate the role of inflammation in 20-HETE regulation of ischemia-induced angiogenesis.

Methods: We first established a mouse hind limb ischemia model for immunocompetent Balb/C mice and immunodeficient NOD-SCID mice by femoral artery ligation. Groups of Balb/C and NOD-SCID mice were administered a 20-HETE synthesis inhibitor, DDMS, or saline as a solvent control. …

Cytotoxic And Antimicrobial Effects Of Silver-Containing Surfaces, Sarah Goderecci

Theses and Dissertations

This study examines applications of sputtered silver coatings as alternatives to traditional antibiotic treatments. Given the increase in reports of antibiotic-resistant bacteria, new treatments and coatings for in-dwelling medical devices such as catheters and orthopedic implants are necessary. Silver oxide films were deposited onto Ti surfaces to examine the efficacy of such coatings against a variety of bacterial species both in vitro and in vivo. Bacterial growth studies showed that coatings exhibited antimicrobial activity against a range of bacterial species acting either in a bacteriostatic or bactericidal mechanism, depending on the target. Limited toxicity to in vitro mammalian cells was …