Biochemistry Commons^™

Preparation And Crystal Structures Of Chiral And Non-Chiral Mixed Ligand Copper Complexes Containing N-Methyl Imidazole And Various N-Phthaloylalanines, Hiral Patel

Theses, Dissertations and Culminating Projects

A quasiracemate is the cocrystallization of two similarly shaped but chemically distinct molecules of opposite handedness. There has been extensive work done in the field of organic quasiracemates by Dr. Kraig A. Wheeler. Limited work has been done in inorganic quasiracemates. The goal of this study was to prepare various metal based enantiomers, racemates, and quasiracemates using A-phthaloylalanines, and TV-methylimidazole as ligands bound to copper arranged in a centrosymmetric pattern. In this study, we developed a method to prepare A-phthaloylalanines and its copper complexes. Three complexes have yielded crystal structures: a racemate, bis(7V-methylimidazole) bis(D,L-3-chloro-7V-phthaloylalanato)copper(II); a quasiracemate, bis(A-methyhmidazole)(L-3-chloro-iV-phthaloylalanato)(D-3-methyl-7V-phthaloylalanato) copper(II); and an …

Single Cell Analysis For The Characterization Of Cell Populations Using A Live Cell Array, Maureen Ann Walling

Legacy Theses & Dissertations (2009 - 2024)

In the past decade, the shift from whole cell population analyses towards single cell measurement methods and techniques is based on experimental results that reveal significant levels of non-genetic heterogeneity in clonal cell populations. This heterogeneity manifests in multiple aspects of cell activity and is, in part, a result of stochastic noise in processes leading to gene expression, namely transcription and translation. The growing understanding of this occurrence has led to the development of methods to monitor and analyze heterogeneity for a more thorough description of cell populations and overall activity.

Structural Interactions And Dynamics Of Disease Related Proteins By Using Nmr Spectroscopy, Shadakshara Swamy Puttamadappa

Legacy Theses & Dissertations (2009 - 2024)

Nuclear Magnetic Resonance (NMR) is a powerful spectroscopic technique to study the structure, molecular interactions, and dynamics of proteins. Modern NMR instrumentation, advancements in experimental techniques and revolutionary developments in recombinant DNA technology have made NMR a versatile and very convenient tool for biomolecule characterization.

In-Cell And In Vitro Studies Of Disease Related Protein-Protein Interactions Using Nmr-Spectroscopy, Andres Yudiel Maldonado

Legacy Theses & Dissertations (2009 - 2024)

The receptor for advanced glycation end products (RAGE) is a multiligand cell surface macromolecule that plays a central role in the etiology of diabetes, inflammation, and neurodegeneration. The cytoplasmic domain of RAGE, ctRAGE, is critical for RAGE-dependent signal transduction. As the most membrane proximal event, mDia1 binds to ctRAGE and is essential for RAGE ligand-stimulated phosphorylation of AKT and cell proliferation/migration. We show that ctRAGE contains an unusual alpha-turn that mediates the mDia1-ctRAGE interaction and is required for RAGE dependent signaling. The results establish a novel mechanism through which an extracellular signal initiated by RAGE ligands regulates RAGE signaling in …

Advanced Statistical Methods For Biochemical And Forensic Applications Of Raman Spectroscopy, Vitali Sikirzhytski

Legacy Theses & Dissertations (2009 - 2024)

This work is dedicated to the development and application of advanced statistical methods for biochemical and forensic applications of Raman spectroscopy. We developed a new concept of multidimensional Raman signatures for forensic identification of body fluid traces, which was successfully applied to the study of dried blood, semen, saliva, sweat and vaginal fluid stains. Several alternative advanced chemometric techniques were also successfully tested for the problems of pure body fluids discrimination and mixture analysis. Studies of body fluid mixtures with low blood and semen concentrations indicated that the detection limit of a minor contributor was as low as a few …

Homologous And Heterologous Crystallin Protein Interactions As The Molecular Basis Of Inherited Cataract, Priya Ranjan Banerjee

Legacy Theses & Dissertations (2009 - 2024)

Age-related cataract is the most common cause of blindness worldwide. Nearly fifty percent of Americans above the age of 75 are diagnosed with this disease (http://www.nei.nih.gov/eyedata/pbd_tables.asp), and surgical intervention is the sole method of treatment at present (http://www.nei.nih.gov/healthyvision/objective/cataracts.asp). In the developing world, even this treatment is not readily available. These are compelling reasons to search for better treatments to delay, prevent or arrest cataract formation. Recent evidence suggests that age-related cataracts also have a genetic component. Therefore, determining the mechanisms underlying genetic cataracts with a known association to a protein-mutation is one important strategy towards understanding the molecular basis for …

Biophysical Investigations Of The Molecular Basis Of Cataract Associated With The R76s Mutation In Human Gammad-Crystallin, Vurghun Ahmadov

Legacy Theses & Dissertations (2009 - 2024)

Cataract disease results when the eye lens becomes opaque and scatters a significant part of the incoming light into the eye. The lens contains very high concentrations of the lens proteins, called crystallins, which are present at concentrations comparable to those found in protein crystals (about 400-600 mg/mL). Chemical modifications of the crystallins, such as oxidation and deamidation, or genetic mutations are known to result in increased light-scattering in vitro, and are implicated in cataract formation in vivo. Here we present the in vitro work on a mutant protein of human gammaD crystallin (HGD), namely R76S (i.e. Arg 76 to …

Fabrication And Characterization Of Mesoscale Protein Patterns Using Atomic Force Microscopy (Afm), Pei Gao

University of Kentucky Doctoral Dissertations

A versatile AFM local oxidation lithography was developed for fabricating clean protein patterns ranging from nanometer to sub-millimeter scale on octadecyltrichlorosilane (OTS) layer of Si (100) wafer. This protein patterning method can generate bio-active protein pattern with a clean background without the need of the anti-fouling the surface or repetitive rinsing.

As a model system, lysozyme protein patterns were investigated through their binding reactions with antibodies and aptamers by AFM. Polyclonal anti-lysozyme antibodies and anti-lysozyme aptamer are found to preferentially bind to the lysozyme molecules on the edge of a protein pattern before their binding to the interior ones. It …

Changes In Apparent Molar Water Volume And Dkp Solubility Yield Insights On The Hofmeister Effect, Daryl K. Eggers, A. Y. Payumo, R. M. Huijon, D. D. Mansfield, L. M. Belk, A. K. Bui, A. E. Knight

Faculty Publications, Chemistry

This study examines the properties of a 4 × 2 matrix of aqueous cations and anions at concentrations up to 8.0 M. The apparent molar water volume, as calculated by subtracting the mass and volume of the ions from the corresponding solution density, was found to exceed the molar volume of ice in many concentrated electrolyte solutions, underscoring the nonideal behavior of these systems. The solvent properties of water were also analyzed by measuring the solubility of diketopiperazine (DKP) in 2.000 M salt solutions prepared from the same ion combinations. Solution rankings for DKP solubility were found to parallel the …

Lead Discovery And Optimization Strategies Towards The Development Of 4(1h)-Quinolones And 1,2,3,4-Tetrahydroacridone Analogs With Antimalarial Activity, Richard Matthew Cross

USF Tampa Graduate Theses and Dissertations

The goal of our research endeavor was to successfully employ modern lead discovery and optimization strategies towards the development and identification of compounds possessing antimalarial activity. Preliminary data from in vitro screening at the Walter Reed Army Institute of Research identified several chemotypes including 4(1H)-quinolones and 1,2,3,4-tetrahydroacridones to have potent antimalarial activities. Multiple synthetic routes were devised and implemented which enabled the rapid preparation and isolation of over 400 structurally diverse 4(1H)-quinolones and 1,2,3,4-tetrahydroacridones.

Our research towards discovering and optimizing antimalarials was inspired from the severe impact malaria has had on our planet especially on impoverished countries. There are over …

Computational Network Analysis Of The Anatomical And Genetic Organizations In The Mouse Brain, Shuiwang Ji

Computer Science Faculty Publications

Motivation: The mammalian central nervous system (CNS) generates high-level behavior and cognitive functions. Elucidating the anatomical and genetic organizations in the CNS is a key step toward understanding the functional brain circuitry. The CNS contains an enormous number of cell types, each with unique gene expression patterns. Therefore, it is of central importance to capture the spatial expression patterns in the brain. Currently, genome-wide atlas of spatial expression patterns in the mouse brain has been made available, and the data are in the form of aligned 3D data arrays. The sheer volume and complexity of these data pose significant challenges …

An Upper Limit For Macromolecular Crowding Effects, Andrew C. Miklos, Congang Li, Courtney D. Sorell, L. Andrew Lyon, Gary J. Pielak

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Background: Solutions containing high macromolecule concentrations are predicted to affect a number of protein properties compared to those properties in dilute solution. In cells, these macromolecular crowders have a large range of sizes and can occupy 30% or more of the available volume. We chose to study the stability and ps-ns internal dynamics of a globular protein whose radius is similar to 2 nm when crowded by a synthetic microgel composed of poly(N-isopropylacrylamide-co-acrylic acid) with particle radii of similar to 300 nm.

Results: Our studies revealed no change in protein rotational or ps-ns backbone dynamics and only mild …

Strategies To Resolve The Three-Dimensional Structure Of The Genome Of Small Single-Stranded Icosahedral Viruses, Eduardo Sanz Garcia

Theses and Dissertations

The aim of this study is the three-dimensional structural characterization of the genome packaging inside viral capsids via cryo-electron microscopy and three-dimensional reconstruction. The genome of some single-stranded viruses can be densely packaged within their capsid shells. Several stretches of the genome are known to adopt stable secondary structures, however, to date, little is known about the three-dimensional organization of the genome inside their capsid shells. Two techniques have been developed to facilitate the structural elucidation of genome packaging: the asymmetric random-model method, and the symmetry-mismatch, random model method. Both techniques were successfully tested with model and experimental data. The …

Chemical Vapor Deposition Of Silanes And Patterning On Silicon, Feng Zhang

Theses and Dissertations

Self assembled monolayers (SAMs) are widely used for surface modification. Alkylsilane monolayers are one of the most widely deposited and studied SAMs. My work focuses on the preparation, patterning, and application of alkysilane monolayers. 3-aminopropyltriethoxysilane (APTES) is one of the most popular silanes used to make active surfaces for surface modification. To possibly improve the surface physical properties and increase options for processing this material, I prepared and studied a series of amino silane surfaces on silicon/silicon dioxide from APTES and two other related silanes by chemical vapor deposition (CVD). I also explored CVD of 3-mercaptopropyltrimethoxysilane on silicon and quartz. …

Dual Inhibition Of Sodium-Mediated Proton And Calcium Efflux Triggers Non-Apoptotic Cell Death In Malignant Gliomas, William Harley, Candace Floyd, Tamara Dunn, Xiao-Dong Zhang, Tsung-Yu Chen, Manu Hegde, Hasan Palandoken, Michael H. Nantz, Leonardo Leon, K.L. Carraway Iii, Bruce Lyeth, Fredric A. Gorin

Chemistry and Biochemistry

Malignant glioma cells maintain an elevated intracellular pH (pH_i) within hypoxic–ischemic tumormicroenvironments through persistent activation of sodium–proton transport (McLean et al., 2000). Amiloride has been reported to selectively kill human malignant glioma cell lines but not primary astrocytes (Hegde et al., 2004). While amiloride reduces pH_i of malignant gliomas by inhibiting isoform 1 of sodium–proton exchange (NHE1), direct acidification was shown to be cytostatic rather than cytotoxic. At cytotoxic concentrations, amiloride has multiple drug targets including inhibition of NHE1 and sodium–calciumexchange. Amiloride's glioma cytotoxicity can be explained, at least in part, by dual inhibition of NHE1 and …

Measurements Of Gas‐Phase Inorganic And Organic Acids From Biomass Fires By Negative‐Ion Proton‐Transfer Chemical‐Ionization Mass Spectrometry, P. Veres, James M. Roberts, I. R. Burling, C. Warneke, Joost De Gouw, Robert J. Yokelson

Chemistry and Biochemistry Faculty Publications

[1] Emissions from 34 laboratory biomass fires were investigated at the combustion facility of the U.S. Department of Agriculture Fire Sciences Laboratory in Missoula, Montana. Gas-phase organic and inorganic acids were quantified using negative-ion proton-transfer chemical-ionization mass spectrometry (NI-PT-CIMS), open-path Fourier transform infrared spectroscopy (OP-FTIR), and proton-transfer-reaction mass spectrometry (PTR-MS). NI-PT-CIMS is a novel technique that measures the mass-to-charge ratio (m/z) of ions generated from reactions of acetate (CH₃C(O)O⁻) ions with inorganic and organic acids. The emission ratios for various important reactive acids with respect to CO were determined. Emission ratios for isocyanic acid (HNCO), 1,2 …

High Flow Air Sampling For Field Detection Using Gas Chromatography-Mass Spectrometry, Jacolin Ann Murray

Theses and Dissertations

The ability to rapidly detect and identify hazardous analytes in the field has become increasingly important. One of the most important analytical detection methods in the field is gas chromatography-mass spectrometry (GC-MS). In this work, a hand-portable GC-MS system is described that contains a miniature toroidal ion trap mass analyzer and a low thermal mass GC. The system is self-contained within the dimensions of 47 x 36 x 18 cm and weighs less than 13 kg. Because the instrument has a small footprint, it was used as the detector for an automated near-real-time permeation testing system. In permeation testing, materials …

Kinetic, Mechanistic, And Structural Investigation Of Features Controlling Stereoselectivity Of (R)- And (S)-Hydroxypropyl Com Dehydrogenases From Xanthobacter Autrophicus Strain Py2, Dariusz Adam Sliwa

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Enantiopure alcohols are valuable intermediates in fine organic synthesis, in particular for preparation of biologically active compounds. The necessity of preparing single enantiomer drugs in an optically pure form has triggered much research, especially in the pharmaceutical industry. The biocatalytical production of chiral alcohols by alcohol dehydrogenase enzymes is characterized by the asymmetric reduction of the corresponding ketones, usually with high degree of stereoselectivity. The commercial value of the enzymes as stereoselective biocatalysts has been a significant driving force in understanding features that control their mechanism of catalysis and stereoselectivity. This work focuses on two enantiocomplementary dehydrogenase enzymes ((R …

Systematic Approach To Electrostatically Induced 2d Crystallization Of Nanoparticles At Liquid Interfaces, Sumit Kewalramani, Suntao Wang, Yuan Lin, Huong Giang Nguyen, Qian Wang, Masafumi Fukuto, Lin Yang

Faculty Publications

We report an experimental demonstration of a strategy for inducing two-dimensional (2D)crystallization of charged nanoparticles on oppositely charged fluid interfaces. This strategy aims to maximize the interfacial adsorption of nanoparticles, and hence their lateral packing density, by utilizing a combination of weakly charged particles and a high surface charge density on the planar interface. In order to test this approach, we investigated the assembly of cowpea mosaic virus (CPMV) on positively charged lipid monolayers at the aqueous solution surface, by means of in situX-ray scattering measurements at the liquid–vapor interface. Theassembly was studied as a function of the solution …

Work Towards The Isolation And Characterization Of The Muscle Isoform Of Glucose 1,6-Bisphosphatase, Caleb J. Hiller

Theses and Dissertations

Glucose 1,6-bisphosphate is an important small molecule involved in the regulation of glycolysis. Four enzymes synthesize this compound. One enzyme is known to degrade it, glucose 1,6-bisphosphatase. Other groups have produced work that indicates that there are two isoforms of this enzyme, one predominant in the brain and one in the muscle. This thesis contains the work performed in attempts to isolate and characterize the muscle isoform of glucose 1,6-bisphosphatase. While this enzyme was not isolated, much was learned about it and the results from this work may help in the future identification of this enzyme.

Dnmt1 Stability Is Regulated By Proteins Coordinating Deubiquitination And Acetylation-Driven Ubiquitination, Zhanwen Du, Jing Song, Yong Wang, Yiqing Zhao, Kishore Guda, Shuming Yang, Hung Ying Kao, Yan Xu, Joseph Willis, Sanford D. Markowitz, David Sedwick, Robert M. Ewing, Zhenghe Wang

Chemistry Faculty Publications

DNA methyltransferase 1 (DNMT1) is the primary enzyme that maintains DNA methylation. We describe a previously unknown mode of regulation of DNMT1 protein stability through the coordinated action of an array of DNMT1-associated proteins. DNMT1 was destabilized by acetylation by the acetyltransferase Tip60, which triggered ubiquitination by the E3 ligase UHRF1, thereby targeting DNMT1 for proteasomal degradation. In contrast, DNMT1 was stabilized by histone deacetylase 1 (HDAC1) and the deubiquitinase HAUSP (herpes virus–associated ubiquitin-specific protease). Analysis of the abundance of DNMT1 and Tip60, as well as the association between HAUSP and DNMT1, suggested that during the cell cycle the initiation …

Halo Ion Trap Mass Spectrometry: Design, Instrumentation, And Performance, Miao Wang

Theses and Dissertations

New ion trap mass spectrometry (ITMS) instrumentation, the toroidal IT and halo IT, were developed to meet the significant growth in on-site analysis applications. The miniature toroidal IT mass analyzer was operated with radio frequency (RF) trapping voltages of 3 kVp-p or less. Despite its reduced dimensions, it has roughly the same ion trapping capacity as conventional 3D quadrupole ITs. Unit-mass resolved spectra for n-butylbenzene, xenon, and naphthalene were obtained. The desired linear mass scale was obtained using conventional mass-selective instability scan combined with resonance ejection. The halo IT was also based on toroidal trapping geometry and microfabrication technology, consisting …

(1e,3e)-1,4-Bis(4-Methoxyphenyl)Buta1,3-Diene, Gopinathan Narayan, Nigam Rath, Suresh Das

Chemistry & Biochemistry Faculty Works

The title compound, C18H18O2, which exhibits blue emission in the solid state, is an inter­mediate in the preparation of liquid crystals and polymers. The mol­ecule is located on an inversion centre. In the crystal, mol­ecules are arranged in a herringbone motif.

Functionalization Of Diamond And Its Application In High Performance Liquid Chromatography And Solid Phase Extraction, Gaurav Saini

Theses and Dissertations

The primary focus of my work was to chemically functionalize diamond as normal and reversed phases for solid phase extraction (SPE) and high performance liquid chromatography (HPLC). Diamond was functionalized with -NH2 groups via self-limiting adsorption of an amine-containing polymer, polyallylamine (PAAm), onto oxidized diamond particles. The chemical stability of these particles was improved by thermal curing or chemical crosslinking with 1,2,5,6-diepoxycyclooctane. The reversed phase material for SPE was synthesized by reacting amine-functionalized diamond particles with long chain alkyl and a perfluorinated isocyanate. Unlike commercially available silica, functionalized diamond particles were stable under extreme pH conditions (pH 0-14). Functionalized diamond …

Solving The Low Dimensional Smoluchowski Equation With A Singular Value Basis Set, Gregory E. Scott, Martin Gruebele

Chemistry and Biochemistry

Reaction kinetics on free energy surfaces with small activation barriers can be computed directly with the Smoluchowski equation. The procedure is computationally expensive even in a few dimensions. We present a propagation method that considerably reduces computational time for a particular class of problems: when the free energy surface suddenly switches by a small amount, and the probability distribution relaxes to a new equilibrium value. This case describes relaxation experiments. To achieve efficient solution, we expand the density matrix in a basis set obtained by singular value decomposition of equilibrium density matrices. Grid size during propagation is reduced from (100–1000) …

Characterization Of Metastasis-Associated Cell Surface Glycoproteins In Prostate Cancer, Lifang Yang

Theses and Dissertations in Biomedical Sciences

Prostate cancer (PCa) is a major health problem in males in the United States. Its lethality is mostly attributed to the primary tumor metastasizing to distant sites that are highly resistant to conventional therapies. Serum Prostate Specific Antigen (PSA) is the only protein biomarker used in clinic for prediction of prostate cancer recurrence following local therapies. Nonetheless, PSA lacks the ability to predict the behavior of an individual tumor in an individual patient. Therefore, development of reliable biomarkers for detection of metastatic potential in primary tumors, as well as discovery of new therapeutic targets, is in a great need for …

Phosphorus Uptake In Rhodomonas Salina (Wislouch) And Its Effect On Allocation And Elimination In Acartia Tonsa (Dana), Danna Palladino

OES Theses and Dissertations

Phosphorus is a key element in important biochemical compounds, such as RNA and phospholipids, and can become limiting in a variety of marine systems. The uptake of phosphorus into biochemical fractions (protein, low molecular weight (LMW) compounds, lipid, polysaccharide and nucleic acid) in Acartia tonsa fed ³³P -labeled Rhodomonas salina was examined. R. salina was cultured on two variations of one media that in one case contained phosphorus in balance and the other out of balance with relation to other standard f/2 components. The P-balanced (PB) media had a N:P ratio of 24.5, which is higher than that found …

Synthesis And Structural Characterization Of The Peptide Epitope Of The Ovarian Cancer Biomarker Ca125 (Muc16), Rebecca J. Whelan, Zach T. Berman, Lee J. Moore, Kathleen E. Knudson

Faculty & Staff Scholarship

A highly conserved region of 21 amino acids flanked by cysteine residues, contained within a larger repeated domain, has been proposed to be the antibody-binding site in the ovarian cancer biomarker CA125 (MUC16). In this study solid-phase peptide synthesis with Fmoc protection chemistry was used to assemble a 21-mer peptide corresponding to the most frequently occurring antibody binding sequence in CA125. Potentially significant sequence variants were also synthesized. Peptide secondary structure was investigated using Fourier transform infrared spectroscopy, revealing the consensus sequence peptide to be largely unstructured at physiological pH whether the cysteine residues were reduced or were oxidized to …

Axial Temperature Gradients In Gas Chromatography, Jesse Alberto Contreras

Theses and Dissertations

The easiest and most effective way to influence the separation process in gas chromatography (GC) is achieved by controlling the temperature of the chromatographic column. In conventional GC, the temperature along the length of the column is constant at any given time, T(t). In my research, I investigated the effects of temperature gradients on GC separations as a function of time and position, T(t,x), along the column. This separation mode is called thermal gradient GC (TGGC). The research reported in this dissertation highlights the fundamental principles of axial temperature gradients and the separation potential of the TGGC technique. These goals …

Negative Dielectrophoretic Capture Of Bacterial Spores In Food Matrices, Mehti Koklu, Seungkyung Park, Suresh D. Pillai, Ali Beskok

Mechanical & Aerospace Engineering Faculty Publications

A microfluidic device with planar square electrodes is developed for capturing particles from high conductivity media using negative dielectrophoresis (n-DEP). Specifically, Bacillus subtilis and Clostridium sporogenes spores, and polystyrene particles are tested in NaCl solution (0.05 and 0.225 S/m), apple juice (0.225 S/m), and milk (0.525 S/m). Depending on the conductivity of the medium, the Joule heating produces electrothermal flow (ETF), which continuously circulates and transports the particles to the DEP capture sites. Combination of the ETF and n-DEP results in different particle capture efficiencies as a function of the conductivity. Utilizing 20 μm height DEP chambers, “almost complete” and …