Life Sciences Commons^™

Studies Into The Allosteric Regulation Of Adp-Glucose Pyrophosphorylases, Benjamin Luke Hill

Dissertations

The synthesis of glycogen in bacteria and starch in plants is allosterically controlled by the production of ADP-glucose by ADP-glucose pyrophosphorylase. Using computational studies, site directed mutagenesis, and kinetic characterization, and protein crystallography we found a critical region for transmitting the allosteric signal in the Escherichia coli and A. tumefaciens ADP-glucose pyrophosphorylase. Molecular dynamics simulations and structural comparisons with other ADP-glucose pyrophosphorylases provided information to hypothesize communication pathways that link allosteric and active sites, and this was tested by site-directed mutagenesis and kinetic characterization of the mutant enzymes. In addition, the application of x-ray crystallography enabled the pinpointing of the …

Development Of "Inside-Out" Pegylated Crosslinked Hemoglobin Polymers: Novel Hemoglobin-Based Oxygen Carriers (Hboc), Kyle Douglas Webster

Dissertations

The development of an effective blood substitute is urgent due to increasingly common blood shortages, the need to type-match donated blood, and communicable diseases (e.g. HIV) posing risks for blood transfusions around the world. There have been many attempts at creating hemoglobin-based oxygen carriers (HBOC) using a variety of techniques centered around the use of polyethylene glycol (PEG) conjugated to hemoglobin (Hb) tetramers. A novel method, “Inside-Out” PEGylation, has been developed by our lab to produce a polyethylene glycol crosslinked hemoglobin (PEG XL-Hb) polymer. This method utilizes a single PEG backbone that is surrounded by multiple proteins, instead of covering …

Mechanistic Variations Of The Bronsted Linear Free Energy Relationships For Nonezymatic Nucleotidyl Transfer Reactions, Zheng Zhang

Dissertations

As the essential enzymes in human bodies, DNA polymerases play a significant role in DNA replication, repair, genetic recombination, and reverse transcription. In 1956, the enzyme of DNA polymerase I, also named as Pol I, was discovered by Arthur Kornberg and colleagues. Subsequently, the Noble Committee had decided that the Noble Prize in Physiology or Medicine for 1959 was to be awarded to Kornberg for his excellent original work that describes the DNA replication process whereby the DNA polymerase copies the nucleotide sequence of a DNA template strand. Because of the complex enzyme structure in the DNA polymerase, it is …

An Analytical Method For Detecting Toxic Metal Cations Using Cyclotriveratrylene Derivative Capped Gold Nanoparticles, Zachary R. Osner, Richard C. Holz, Daniel P. Becker Ph.D.

Chemistry: Faculty Publications and Other Works

Cyclotriveratrylene-oxime (CTV-oxime) derivatives that terminate with a dithiolate linker were synthesized enabling the supramolecular scaffold to adhere to gold nanoparticles (AuNPs) with the bowl-shaped cavity of the CTV scaffold exposed for utilization in host–guest chemistry. Exposure of these CTV functionalized AuNPs to varying concentrations of di- and trivalent metal cations resulted in the formation of large CTV-AuNP polymeric clusters and an accompanying a shift in the plasmon resonance. These interactions between the CTV-AuNPs and the metal cations in solution provides proof-of-concept that supramolecular functionalized AuNPs can be used as a simple and straightforward, on-site detection system for toxic metal cations …

Metal-Free Tandem Beckmann–Electrophilic Aromatic Substitution Cascade Affording Diaryl Imines, Ketones, Amines, And Quinazolines, Samuel Sarsah, Marlon R. Lutz Jr., Kailyn Chichi Bobb, Daniel Becker

Chemistry: Faculty Publications and Other Works

A cascade reaction sequence involving a Beckmann rearrangement on benzophenone oxime followed by an electrophilic aromatic substitution (EAS) on the intermediate nitrilium ion affords N-phenyl diaryl imines that may then be hydrolyzed to ketones, or reduced to the corresponding amines. Reaction with benzonitrile afforded 2,4-diphenylquinazoline through a Beckmann–Ritter–EAS cascade.

Study Of Escherichia Coli Adp-Glucose Pyrophosphorylase Catalysis: Investigating Critical Roles Of Conserved Arg32 And Lys42 Residues, Angela Lynn Mahaffey

Dissertations

Carbohydrates have been most notable as energy sources for mammals, bacteria (glycogen) and plants (starch) – and in many other species. As such the biosynthesis of carbohydrates is essential to the sustainability of many forms of life, on earth. Adenosine-‘5-diphosphate glucose pyrophosphate (ADP-Glc pyrophosphorylase; ADP-Glc PPase) is the allosterically controlled “first committed step” in both the biosynthetic pathways of starch (~25% amylose and ~75% amylopectin, in plants and algae) and glycogen (in bacteria), preceding the starch/glycogen synthase reaction. By catalyzing the following reaction, ATP + α –D-Glc-1P ADP-Glc + PPi , ADP-Glc PPase functions as the primary enzyme in the …

Structure And Molecular Mechanism Of A Plp/Gaba Dependent Transcription Regulator Gabr, Rui Wu

Dissertations

GabR is a member of the MocR/GabR subfamily of the GntR family of bacterial transcription regulators. It regulates the metabolism of γ-aminobutyric acid (GABA), an important nitrogen and carbon source in many bacteria. The crystal structures reported here show that this protein has evolved from the fusion of a type I aminotransferase and a winged helix-turn-helix (wHTH) DNA binding protein to form a chimeric protein that adopts a dimeric head-to-tail configuration. The pyridoxal 5′-phosphate (PLP)-binding regulatory domain of GabR is therefore an example of a coenzyme playing a role in transcription regulation rather than in enzymatic catalysis. Our structural and …

Mutations In Phospholamban Alter The Structure And Function Of The Calcium Atpase Regulatory Complex, Neha Abrol

Dissertations

Phospholamban (PLB) is an integral membrane protein that plays an important role in regulation of cardiac calcium handling and contractility. PLB exists as a homopentamer in the membrane, which upon deoligomerization into active monomers reversibly inhibits sarco/endoplasmic reticulum calcium ATPase (SERCA). Mutations in PLB that change the PLB monomer-pentamer equilibrium result in dysregulation of SERCA. To determine the structural and regulatory role of the C-terminal residues of PLB in the membranes of living cells, we fused fluorescent protein tags to PLB and SERCA. We then studied the effect of C-terminal alanine substitutions and truncation mutations on PLB oligomerization and SERCA …

Catalytic Mechanism And Maturation Of The Metalloenzyme Nitrile Hydratase, Natalie I. Gumataotao

Dissertations

Nitrile hydratases are metalloenzymes that catalyze the hydration of nitriles to their corresponding amides in a specific manner at ambient pressures and temperatures at neutral pH. Traditional industrial methods require high temperature and pressure, extreme pH, and heavy metals. NHases are used as biocatalysts in the large scale industrial production of amide precursors to textiles, animal feedstock, and polymers. Notably, NHase is used in the production of ~100,000 tons of acrylamide annually by the Mitsubishi Corporation.

Despite being used extensively in industry, questions remain about NHase. The catalytic mechanism is not defined. Understanding the way in which the nitrile is …

The Dape-Encoded N-Succinyl-L,L-Diaminopimelic Acid Desuccinylase (Dape) From Haemophilus Influenzae As A Prospective Target For The Development Of Novel Antibiotics, Anna Starus

Dissertations

The rapid emergence of bacteria that are resistant to todayfs antibiotics makes them more and more ineffective. Consequently, the need for a novel class of antibacterial agents is rapidly increasing. The importance of this project is emphasized by the emergence of several pathogenic bacterial strains that are resistant to all currently available antibiotics on the market today. One way to approach this problem is to develop drugs that target essential bacterial biosynthetic pathways. Based on bacterial genetic information, the meso]diaminopimelate (mDAP)/lysine biosynthetic pathway offers several potential anti]bacterial targets that have not been yet explored. One of these, the dapE-encoded N-succinyl-L,L-diaminopimelic …

Lysine Biosynthesis In Bacteria: A Metallodesuccinylase As A Potential Antimicrobial Target, Danuta M. Gillner, Daniel P. Becker Ph.D., Richard C. Holz

Chemistry: Faculty Publications and Other Works

In this review, we summarize the recent literature on dapE-encoded N-succinyl-l,l-diaminopimelic acid desuccinylase (DapE) enzymes, with an emphasis on structure–function studies that provide insight into the catalytic mechanism. Crystallographic data have also provided insight into residues that might be involved in substrate and hence inhibitor recognition and binding. These data have led to the design and synthesis of several new DapE inhibitors, which are described along with what is known about how inhibitors interact with the active site of DapE enzymes, including the efficacy of a moderately strong DapE inhibitor.

Critical Functions Specified By The Mll Cxxc Domain Determine Leukemogenic Capacity, Noah Warren Birch

Dissertations

TheMixed Lineage Leukemia(MLL) gene can participate in chromosomal translocations which generate a fusion protein leading to acute leukemia. A better understanding of how MLL fusion proteins contribute to leukemia is necessary in order to develop more effective treatments. In my dissertation project, I investigated the functional role of amino acids within the MLL CXXC domain to determine how specific residues contribute to leukemogenic capacity.

MLL fusion proteins retain the amino-terminal portion of MLL including the CXXC DNA-binding domain while the carboxy-terminal portion is comprised of a fusion partner. The closest homolog of MLL, MLL2 (alternatively named MLL4), also contains a …

Biophysical Characterization Of Tryptophan Locales, Mg²+ Binding And Protein Folding In Gα Subunits, Matthew Najor

Dissertations

The objective of this study is to understand the structure of guanine nucleotide - binding (G) proteins using a variety of spectroscopic tools. G proteins are membrane-bound proteins consisting of α, β, and γ subunits required for the transduction of extracellular signals to various intracellular effectors. Activation of G protein coupled receptors by neurotransmitters or hormones result in a conformational change of a G protein that is triggered by the exchange of guanosine 5'- diphosphate (GDP) bound to the  subunit for guanosine 5'- triphosphate (GTP) and concomitant dissociation of the  dimer.

Wild type (WT) Giα1 has three tryptophan …

Role Of Non-Coding Rna Nc4 In Mll And Cyp33 Mediated Regulation Of Hoxc8, Jessica Arvindbhai Solanki

Dissertations

MLL or Mixed Lineage Leukemia gene is clinically known for its involvement in genetic translocation with more than 70 different partners identified each giving rise to a highly leukemogenic fusion protein. With a poor prognosis of MLL related leukemia, an investigation into its role during hematopoiesis has been a very active field of research. In order to design strategies to combat the MLL leukemia, it becomes essential to delineate the molecular mechanisms behind the function of MLL wild type protein. Wild type MLL is a transcriptional maintenance protein from the Trithorax Group (TrxG) that resides in complex with other chromatin …

Polyomavirus Enhancer Activator 3 (Pea3), A Member Of The Ets Family Of Transcription Factors, Is A Transcriptional Activator Of Notch-1 And Notch-4 In Breast Cancer: An Opportunity For Novel Combinational Therapy, Anthony George Clementz

Dissertations

Women diagnosed with triple-negative breast cancer have the worst overall prognosis and frequently present with metastatic tumors. To date, there are no targeted therapies available to combat this aggressive form of breast cancer due to the lack of expression of well-known targets such as ER-alpha, PR, or HER2/neu. Therefore, there is an immediate need to identify novel targets that are responsible for the proliferation, survival, and invasive phenotype. Notch-1 and Notch-4, both potent breast oncogenes, are overexpressed in triple-negative breast cancers-associated with the poorest overall survival. PEA3 (polyomavirus enhancer activator 3), a member of the Ets family of transcription factors, …

New Cyclophanes As Supramolecular Scaffolds: The Synthesis Of Tribenzo-1,4,7-Triazacyclononatriene., Andria M. Panagopoulos

Dissertations

Supramolecular chemistry involves the formation of complex molecular entities that have the capacity to participate in specific molecular recognition of guest molecules. A commonly employed scaffold in supramolecular chemistry is the trimeric crown-shaped molecule cyclotriveratrylene (CTV). CTV has been studied extensively for its capability of binding a number of smaller organic and organometallic guests within its bowl-shaped cleft and has been used as a building block enabling the construction of more complex cryptophanes. The goal of this research is the synthesis and characterization of a novel cyclophane, tribenzo-1,4,7-triazacyclononene and derivatives thereof. These new cyclophanes should have greater versatility than the …

Structure-Function Relationship Studies Of The Udp-Glucose Pyrophosphorylase From Escherichia Coli, Agnieszka Maria Orlof

Master's Theses

UDP-Glucose Pyrophosphorylase (UDP-Glc PPase) is a key enzyme of the carbohydrate metabolic pathway widely used among prokaryotes and eukaryotes. In plants, UDP-Glc PPase is necessary for sucrose synthesis while mammals utilize this enzyme for the production of glycogen. A bacterium such as Escherichia coli uses UDP-Glc PPase for biosynthesis of the lipopolysscharide core which forms a cell wall.

This study focused on UDP-Glc PPase from Escherichia coli which is encoded by both galU and galF genes. The first part of this project investigated amino acids that could play an important role in the function of UDP Glc PPase (GalU). Based …

Singlet-Oxygen-Mediated Inactivation Of Acetylcholinesterase: A Comparison Of Purified Enzyme In Solution And Enzyme Bound To K562 Leukemia Cells, Gail D. Deadwyler

Dissertations

No abstract provided.

The Effects Of Ethanol On The Developing Serotonergic Neurons, Jung-Ae Kim

Dissertations

No abstract provided.

Cloning Of Multiple Novel Human Trinucleotide Repeat Containing Cdna's: A Novel Application Of Rapid Amplification Of Cdna Ends (Race), James P. Carney

Dissertations

No abstract provided.

The Endocytosis Of Iodinated Protein Substrates In Cultured Human Skin Fibroblasts, Daniel P. Wilcockson

Master's Theses

No abstract provided.

Transformation Related Defects In Vacuolar Acidification And Mannose 6-Phosphate Receptor Function: Effects On Lysosomal Targeting, Valerie Pracht

Dissertations

No abstract provided.

Lysosomal Enzyme Secretion In Tumor Cells And The Purification And Partial Characterization Of A 67 Kda Cathepsin B-Like Enzyme Secreted By B16 Melanoma Cells, Charles Achkar

Dissertations

No abstract provided.

A Proposal For United States/Polish Collaboration On Energy And The Environment, Carole B. Szpunar

Chemistry: Faculty Publications and Other Works

Argonne National Laboratory, a U.S. Department of Energy facility that is operated by the University of Chicago and located southwest of Chicago, Illinois, has proposed a collaborative effort by the United States and Poland to conduct applied research on energy and environmental topics of common interest. Through this collaboration, evaluative studies on energy and environmental assessment issues could be developed and implemented by Argonne and one or more sister institutions in Poland, which could lead to active research, development, and demonstration joint activities. The objective of this initiative is to produce strategies and options for review by Poland's decision makers …

Tryptidase: A Novel Trypsin-Like Enzyme From Rat Skin. Isolation And Characterization, Vincent J. Braganza

Dissertations

No abstract provided.

Role Of The Pseudomonas Aeruginosa Reca Analogue In Lysogeny And Recombination, Tyler Alan. Kokjohn

Dissertations

No abstract provided.

Transition-State Analog Affinity Purification Of Proteases, Arunbhai Patel

Dissertations

No abstract provided.

The Purification, Properties And Chemical Kinetics Of Cathepsin B1, Andrew Stanley Bajkowski

Master's Theses

No abstract provided.

Aldehydic Transition State Analogue Inhibitors Of Α-Chymotrypsin, Alexandra Constance Cheerva

Master's Theses

No abstract provided.

Aluminum Metabolism In Health And Disease, Jay Eliot Gorsky

Dissertations

No abstract provided.