Biochemistry Commons^™

Defining The Role Of Phosphorylation And Dephosphorylation In The Regulation Of Gap Junction Proteins, Hanjun Li

Theses & Dissertations

Gap junctions are intercellular channels that permit the free passage of ions, small metabolites, and signaling molecules between neighboring cells. In the diseased human heart, altered ventricular gap junction organization and connexin expression (i.e., remodeling) are key contributors to rhythm disturbances and contractile dysfunction. Connexin43 (Cx43) is the dominant gap junction protein isoform in the ventricle which is under tight regulation by serine/tyrosine phosphorylation. Phosphorylation and dephosphorylation regulate many aspects of Cx43 function including trafficking, assembly and disassembly, electrical and metabolic coupling at the plaque, as well as to modulate the interaction with other proteins.

Serine phosphorylation has long been …

Regulation Of Alteration/Deficiency In Activation 3 (Ada3) By Acetylation And Its Role In Cell Cycle Regulation And Oncogenesis, Shashank Srivastava

Theses & Dissertations

The ADA3 (Alteration/Deficiency in Activation 3) protein is a transcriptional adaptor protein that was initially discovered as a component of several HAT (Histone Acetyltransferase) complexes, the enzyme complex responsible for histone acetylation, which is a prerequisite for transcription. Earlier the studies from Dr. Band’s laboratory and that of others’ have deciphered a crucial role of ADA3 in cell cycle regulation (both through G₁/S and G₂/M phase transitions) and in maintaining the genomic stability.

While our laboratory investigated the mechanism behind the role of ADA3 in G₁/S transition, the same remained unknown for G_{2 …}

Mitogen And Morphogen Signaling Dysregulation: Pathophysiological Influence In Pancreatic Cancer And Alzheimer’S Disease, Eric Cruz

Theses & Dissertations

Although the etiology of a particular disease will vary, there are genetic and epigenetic bottlenecks that frequently converge resulting in dysregulation of mitogenic and morphogenetic signaling. This propensity is acutely experienced in malignancy and neurodegenerative disease.

Here, we have first investigated the role of dysregulated signaling in the context of pancreatic cancer (PC). Morphogenetic signaling has been regarded as a pleiotropic pathway with the potential to promote and inhibit metastatic features. Our investigation of bone morphogenetic protein 2 (BMP-2), an archetypical member of the BMP superfamily, has revealed the presence of extracellular, intracellular, and long non-coding RNA products. Our findings …

Dna Polymerase Zeta-Dependent Mutagenesis: Molecular Specificity, Extent Of Error-Prone Synthesis, And The Role Of Dntp Pools, Olga V. Kochenova

Theses & Dissertations

Despite multiple DNA repair pathways, DNA lesions can escape repair and compromise normal chromosomal replication, leading to genome instability. Cells utilize specialized low-fidelity Translesion Synthesis (TLS) DNA polymerases to bypass lesions and rescue arrested replication forks. TLS is a highly conserved two-step process that involves insertion of a nucleotide opposite a lesion and extension of the resulting aberrant primer terminus. The first step can be performed by both replicative and TLS DNA polymerases and, because of non-instructive DNA lesions, often results in a nucleotide misincorporation. The second step is almost exclusively catalyzed by DNA polymerase ζ …

Towards Understanding Osmolyte Effects On Folate(S) And Dihydrofolate Reductase Proteins, Purva Prashant Bhojane

Doctoral Dissertations

Osmolytes are small molecules that alter water activity and probe role of water in biological processes. Osmotic stress approach explored the role of water in ligand binding to dihydrofolate reductase (DHFR). DHFR catalyzes NADPH dependent reduction of dihydrofolate (DHF) to tetrahydrofolate (THF), which is essential for the synthesis of DNA, amino acids and other metabolic intermediates. R67 DHFR is a plasmid-encoded DHFR that confers resistance against trimethoprim, which is a potent inhibitor of E.coli chromosomal DHFR.

Osmolytes addition decreases the affinity of the substrate towards both the DHFRs. Weak preferential interactions between the osmolytes and DHF impede substrate binding to …

Electrogenerated Chemiluminescence Study Of Semiconductor Nanoparticles Towards Sensitive Detection Of Biomolecules, Yiliyasi Wusimanjiang

Dissertations

The main focus of this dissertation is to unfold the fundamental aspects of electrogenerated chemiluminescence (ECL) generation from semiconductor nanoparticles (also known as quantum dots or QDs) within different ECL systems. The ECL and photo-physical interactions between the CdTe QDs (λ_emission= ~760 nm) and the CdSe QDs (λ_emission= ~550 nm), as well as the effects of carbon nanotubes on ECL of QDs were separately investigated. Optimum experimental conditions for peptide bond formation on an electrode surface through EDC (1-ethyl-3-(-3-dimethylaminopropyl) carbodiimide hydrochloride)/NHS (N-Hydroxysulfosuccinimide) coupling were also revealed using cyclic voltammetry technique. Based on the information …

Utilization Of Ferrioxamine Microarrays For The Rapid Detection Of Pathogenic Bacteria, Nigam Bir Arora

Open Access Dissertations

Siderophores are low-molecular weight species utilized by bacteria for the sequestration of iron, an essential nutrient. Siderophores and their cognate receptors are considered to be virulence factors, due to their prominent role in pathogenicity. The work presented here focuses on ferrioxamine (FOx) as an “immutable” ligand for pathogen detection. A number of bacterial strains expressing high-affinity FOx receptors were identified by a proteomic BLAST search, and screened against microarrays patterned with FOx conjugates for detection using label-free optical imaging. Aspects such as inkjet printing and surface chemistry, iron-limiting conditions and bacterial selection protocols, and linker conjugate design were addressed and …

Functional And Structural Characterization Of The Mevalonate Diphosphate Decarboxylase And The Isopentenyl Diphosphate Isomerase From Enterococcus Faecalis, Chun-Liang Chen

Open Access Dissertations

Enterococcus faecalis causes a diverse range of nosocomial infections (in wounds, the gastrointestinal tract, the blood stream and the endocardium), and multidrug-resistant strains have become a serious issue across countries. Vancomycin, a FDA-approved drug for the disruption of the bacterial cell wall biosynthesis, has been utilized to treat infectious diseases caused by Enterococci; however, the prevalence of vancomycin-resistant enterococci (VRE) threatens communities all over the world. We aim at developing novel therapeutic strategies to control bacterial growth of Enterococci, and we focus on targeting two essential enzymes involved in poly-isoprenoid biosynthesis in Enterococcus faecalis; one is the mevalonate diphosphate decarboxylase …

Sugarcane Bagasse Hydrolysis Enhancement Using Bsa, Antonio Carlos Freitas Dos Santos

Open Access Theses

Lignocellulose is composed of polysaccharides linked to lignin and other aromatic compounds, making the sugars not readily available to fermentation. This entails that biomass must go through the unit operations of pretreatment and enzyme hydrolysis. Pretreatment opens the structure to allow the enzymes to act on and hydrolyze cellulose and hemicellulose to glucose and/or xylose which in turn are fermented to ethanol. Concomitantly, the enzymes interact with soluble phenols and insoluble solids derived from lignin that inhibit hydrolysis. This leads to high enzyme loadings and higher production costs. Soluble phenols can be eliminated through washing. Insoluble lignin, however, demands another …

Antibacterial Activity Of Essential Oil Encapsulated Sodium Iota-Carrageenan Fibers, Carlos D. Carter

Open Access Theses

Spoilage microorganisms cause food waste and loss of quality. While the foodborne pathogen outbreaks lead to thousands of hospitalizations and deaths. Essential oils (EOs), plant extracts, possess the required antimicrobial activities and thus their usage stands out as a feasible approach for controlling the undesirable bacterial growth in food systems. However, EOs are highly volatile and lose their activity upon exposure to environmental conditions. In this regard, their encapsulation in Generally Recognized As Safe (GRAS) matrices such as food grade polysaccharides especially iota-carrageenan could be one of the viable alternatives. Iota-carrageenan, sulfated polysaccharide from marine algae, is being used in …

The Role Of Hif1alpha And Hif2alpha In Muscle Development And Satellite Cell Function, Shiqi Yang

Open Access Theses

Hypoxia inducible factors (HIFs) are central mediators of cellular responses to fluctuations of oxygen, an environmental regulator of stem cell activity. Muscle satellite cells are myogenic stem cells whose quiescence, activation, self-renewal and differentiation are influenced by microenvironment oxygen levels. However, the in vivo roles of HIFs in quiescent satellite cells and activated satellite cells (myoblasts) are poorly understood. Expression analyses indicate that HIF1α and HIF2α are preferentially expressed in pre- and post-differentiation myoblasts, respectively. Interestingly, double knockout of HIF1α and HIF2α (HIF1α/2α dKO) in embryonic myoblasts results in apparently normal muscle development and growth. However, HIF1α/2α dKO in postnatal …

Tetrameric Photosystem I: From Initial Discovery And Characterization In Chroococcidiopsis Sp. Ts-821 To Exploration Of Its Distribution And Understanding Of Its Significance In Cyanobacteria, Meng Li

Doctoral Dissertations

Photosystem I (PSI) forms trimeric complexes in most characterized cyanobacteria. We had reported the tetrameric form of PSI in the unicellular cyanobacterium, Chroococcidiopsis sp. TS-821 (TS-821). Using Cryo-EM, a 3D model of the PSI tetramer structure at 11.5 [Angstrom] resolution was obtained and a 2D map within the membrane plane of at 6.1 [Angstrom]. In contrast to the three-fold symmetry in trimeric PSI crystal structure from T. elongatus, two different inter-monomer interactions involving PsaLs are found in the PSI tetramer. Phylogenetic analysis based on PsaL protein sequences shows that TS-821 is closely related to heterocyst-forming cyanobacteria. Additionally, this tetrameric …

Membrane Chromatography For Bioseparations: Ligand Design And Optimization, Zizhao Liu

Graduate Theses and Dissertations

Membrane chromatography, or membrane adsorber, represents an attractive alternative to conventional packed bed chromatography used in downstream processing. Membrane chromatography has many advantages, including high productivity, low buffer consumption and ease to scale up. This doctoral dissertation focuses on developing novel polymeric ligands for protein separations using membrane chromatography. Atom transfer radical polymerization (ATRP), known as a controlled radical polymerization technique, has been used to control the architecture of grafted polymeric ligands. The center theme of this dissertation is to develop new polymeric ligands and investigate how the polymer’s property (e.g. flexibility, hydrophobicity) and architecture (e.g. chain density, chain length) …

Bioorthogonal Reactions: Synthesis And Evaluation Of Different Ligands In Copper Catalyzed Azide-Alkyne1,3-Dipolar Cycloaddition (Cuaac), Zainab Abdullah Almansaf

Graduate Theses and Dissertations

The Copper CatalyzedAzide-Alkyne1,3-Dipolar Cycloaddition (CuAAC) reaction has unique features that qualify it to be one of the best click reactions. Its applications have been shown in different aspects and for multiple purposes. The oxidative degradation of biological systems (labile proteins and live cells) is, however, generally recognized as the major problem when using this reaction in living systems. Reactive oxidation species can be easily produced in the presence of copper(II), ascorbate and air, and this is the main cause of toxicity. However, the uses of ligands have shown a major impact on reducing copper toxicity, protecting Cu(I) from the redox …

The Role Of Phosphorylation In Pam2 Motif-Containing Proteins Mediated Messenger Rna Deadenylation, Kai-Lieh Huang

Dissertations & Theses (Open Access)

Phosphorylation regulates many cellular processes. However, its role in mRNA deadenylation, a process to remove poly adenosines from the mature mRNA 3’ end tail, is unclear. The length of poly(A) tail determines mRNA stability and translation efficiency. Poly(A)-binding protein (PABP), which binds to newly synthesized poly(A) tails homogeneously and is known as a scaffold protein for PAM2 motif-containing proteins, plays a pivotal role in the shortening of poly (A) tails. This study is to examine the role of phosphorylation of PAM2 motif–containing proteins in regulating their interactions with PABP and mRNA deadenylation function.

The PAM2 motif, a region required for …

Engineered Human Acidic Fibroblast Growth Factor (Fgf1) With An Enhanced Thermal And Proteolytic Stability, Duaa Abdullah Almansaf

Graduate Theses and Dissertations

Fibroblast growth factor receptor (FGFR) is made up of three significant domains. The most important domain is the intracellular domain where the dimerization and autophosphorylation occur. Fibroblast growth factor (FGF) interacts with specific FGFR to regulate many cellular processes during the embryonic stage. Furthermore, FGF is significant for adults because FGF plays an important role in regulating cellular differentiation as well as wound healing. The cellular regulating processes are initiated through binding FGF to heparin followed by binding FGF/heparin to FGFR to form FGF/heparin/FGFR complex. Thus, FGFR is dimerized and autophosphorylated. The phosphorylation of FGFR triggers downstream signaling pathways, which …

Interactions In The Cpsrp Dependent Targeting Of Light Harvesting Chlorophyll Binding Protein To The Thylakoid Membrane, Rory Henderson

Graduate Theses and Dissertations

Targeting of proteins is a critical component of cellular function. A universally conserved targeting system of the cytosol utilizes a signal recognition particle (SRP) to target many proteins contranslationally to the endoplasmic reticulum in eukaryotes or the inner membrane in prokaryotes. A homologous SRP system exists in the chloroplast that delivers light harvesting chlorophyll binding proteins (LHCP) to they thylakoid membrane. The chloroplast SRP (cpSRP) is a heterodimer composed of a novel 43 kDa subunit and a 54 kDa subunit homologous to a component of the SRP system, SRP54. Many details regarding the interactions between the proteins of the cpSRP …

Characterization Of The Substrate Interactions And Regulation Of Protein Arginine Methyltransferase, Yalemi Morales

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Protein arginine methylation is a posttranslational modification catalyzed by the family of proteins known as the protein arginine methyltransferases (PRMTs). Thousands of methylated arginines have been found in mammalian cells. Many targets of arginine regulation are involved in important cellular processes like transcription, RNA transport and processing, translation, cellular signaling, and DNA repair. Since PRMT dysregulation has been linked to a variety of disease states, understanding how the activity of the PRMTs is regulated is of paramount importance. PRMT1 is the predominant PRMT, responsible for about 85% of all arginine methylation in cells, but very little is known about how …

Lignin Maximization: Analyzing The Impact Of Different Feedstocks And Feedstock Ratios Using Organosolv Fractionation, Marc Banholzer

Masters Theses

Over-exploitation of fossil fuels coupled with increasing pressure to reduce carbon emissions are prompting a transition from conventional petrochemical feedstocks to sustainable and renewable sourced carbon. The use of lignocellulosic biomass as a feedstock for integrated biorefining is of current high interest, as separation into its component parts affords process streams of cellulose, hemicellulose and lignin, each of which can serve as a starting point for the production of biobased chemicals and fuels. Given the large number of potential sources of lignocellulosic feedstocks, the biorefinery will need to adapt to the supplies available over a normal growing season. Of particular …

Effect Of Varying Rumen Degradable And Undegradable Protein On Milk Production And Nitrogen Efficiency In Lactating Dairy Cows Under Summer Conditions, Jeffrey D. Kaufman

Masters Theses

The objective is to determine the effect of reducing nitrogen input through feeding low rumen degradable protein (RDP) and rumen undegradable protein (RUP) proportions on milk production, nitrogen efficiency and metabolism in heat-stressed cows. Forty-eight mid-lactating, Holstein cows were assigned to treatments using a randomized block design in a 2x2 factorial arrangement of treatments (n = 12/treatment). Treatments included two levels of RDP (10 and 8%) and two levels of RUP (8 and 6%). From d 1 to 21, a common diet (10% RDP-8% RUP) was fed to cows followed with their respective treatment diets fed from d 22 to …

A Novel Role Of Human Dna Damage Checkpoint Protein Atr In Suppressing Ca2+ Overload-Induced Parp1-Mediated Necrosis, Hui Wang-Heaton

Electronic Theses and Dissertations

Ataxia telangiectasia and Rad3-related (ATR) is well known for its regulatory role in DNA damage responses (DDR) as a checkpoint kinase that phosphorylates hundreds of protein substrates. However, its role in cellular non-DNA damage stress responses (NDDR) is unknown. Necrosis is one form of cell death and traditionally has been regarded as a passive and uncontrolled cell death. Recently, evidence has emerged to support the concept that necrosis also may occur in a programmed manner and that PARP1 can be a mediator. Active poly (ADP-ribose) polymerase 1 (PARP1) hydrolyzes nicotinamide adenine dinucleotide (NAD⁺) to produce poly (ADP-ribose) (PAR) …

Influence Of Ph And Acidic Side Chain Charges On The Behavior Of Designed Model Peptides In Lipid Bilayer Membranes, Venkatesan Rajagopalan

Graduate Theses and Dissertations

The molecular properties of transmembrane proteins and their interactions with lipids regulate biological function. Of particular interest are interfacial aromatic residues and charged residues in the core helix whose functions range from stabilizing the native structure to regulating ion channels. This dissertation addresses the pH dependence and influence of potentially negatively charged tyrosine, glutamic acid or aspartic acid side chains. We have employed GWALP23 (acetyl-GGALW5LALALALALALALW19LAGA-amide) as favorable host peptide framework. We have substituted W5 with Tyr (Y5GWALP23) and Leu residues with Glu (L12E, L14E or L16E) or Asp (L14D or L16D), and have incorporated specific 2H-labeled alanine residues within the …

Switchgrass Extractives Have Potential As A Value-Added Antimicrobial Against Plant Pathogens And Foodborne Pathogens, Alexander Ian Bruce

Masters Theses

Panicum virgatum (switchgrass), a perennial grass native to North America, is a leading biomass feedstock candidate for the manufacture of cellulosic ethanol. Switchgrass is considered a viable option for biofuel production due to its cheap production cost and ability to grow on marginal land. Biofuel derived from switchgrass has been shown to be very energy efficient, producing 540% more renewable energy versus nonrenewable energy expended. Switchgrass-derived biofuel is also estimated to have greenhouse gas emissions that are 94% lower than emissions from gasoline (Schmer et al 2008). Biofuels are created through biochemical processes that utilize various enzymes and microorganisms for …

Characterization Of Ranbpm Subcellular Localization And Function In Hdac6 Regulation, Louisa M. Salemi

Electronic Thesis and Dissertation Repository

RanBPM has been shown to interact with numerous proteins implicating it in a variety of cellular processes including apoptosis, transcription regulation, cell migration, adhesion and morphology and has been shown to have tumour suppressive functions. RanPBM has been hypothesized to be a scaffolding protein and is part of a large protein complex termed the CTLH complex. Although the homologous complex in yeast has been demonstrated to have E3 ubiquitin ligase activity, whether the evolutionarily conserved human complex retains this activity remains to be determined. In this work we aim to characterize the regions of RanBPM that regulate its subcellular localization …

The Effects Of Phytohormones And Isoprenoids In Dihydroartemisinin-Induced Dormancy In The Erythrocytic Stages Of Plasmodium Falciparum, Marvin Duvalsaint Duvalsaint

USF Tampa Graduate Theses and Dissertations

Our ability to control malaria has been challenged by increasing antimalarial resistance. Plasmodium falciparum undergoes dormancy in the blood stages which is hypothesized to be a means by which they are able to survive under drug pressure. This helps select for resistant parasites which grow following removal of drug. The mechanisms behind dormancy and the subsequent recrudescence are not fully understood but translating knowledge from related organisms which undergo a similar phenomenon might shed some light. Higher plants utilize dormancy during the early development stages to survive under unfavorable conditions, increasing fitness of the seedling and ensuring viability when this …

Quantitative Proteomic Investigation Of Disease Models Of Type 2 Diabetes, Mark Gabriel Athanason

USF Tampa Graduate Theses and Dissertations

PANcreatic DERived factor (PANDER, FAM3B) is a member of a superfamily of FAM3 proteins that are uniquely structured and strongly expressed from the endocrine pancreas and co-secreted with insulin. Unique animal models available to our lab have indicated that PANDER can induce a selective hepatic insulin resistant (SHIR) phenotype whereby insulin signaling is blunted yet lipogenesis is increased. The complexity of the biological networks involved with this process warranted the logical approach of employing quantitative mass spectrometry based proteomic analysis using stable isotope labeling of amino acids in cell culture (SILAC) to identify the global proteome differences between the PANDER …

Significance Of Pten Phosphorylation And Its Nuclear Function In Lung Cancer, Prerna Malaney

USF Tampa Graduate Theses and Dissertations

Phosphorylation mediated inactivation of PTEN leads to multiple malignancies with increased severity. However, the consequence of such inactivation on downstream functions of PTEN are poorly understood. Therefore, the objective of my thesis is to ascertain the molecular mechanisms by which PTEN phosphorylation drives lung cancer. PTEN phosphorylation at the C-terminal serine/threonine cluster abrogates its tumor suppressor function. Despite the critical role of the PTEN C-tail in regulating its function, the crystal structure of the C-tail remains unknown. Using bioinformatics and structural analysis, I determined that the PTEN C-tail is an intrinsically disordered region and is a hot spot for post-translational …

Development Of In Vivo Systems For Detecting And Studying Ribosome Inhibition By Small Molecules, Shijie Huang

Chemistry and Chemical Biology ETDs

The ribosome is the quintessential antibacterial drug target, with many structurally and mechanistically distinct classes of antibacterial agents acting by inhibiting ribosome function. Detecting and quantifying ribosome inhibition by small molecules and investigating their binding modes and mechanisms of action are critical to antibacterial drug discovery and development efforts. To develop a ribosome inhibition assay that is operationally simple, yet provides direct information on the drug target and the mechanism of action, we have developed engineered E. coli strains harboring an orthogonal ribosome controlled green fluorescent protein reporter that produce fluorescent signal when the O-ribosome is inhibited. As a proof …

Regulation Of Palmitoylation Enzymes And Substrates By Intrinsically Disordered Regions, Krishna D. Reddy

USF Tampa Graduate Theses and Dissertations

Protein palmitoylation refers to the process of adding a 16-carbon saturated fatty acid to the cysteine of a substrate protein, and this can in turn affect the substrate’s localization, stability, folding, and several other processes. This process is catalyzed by a family of 23 mammalian protein acyltransferases (PATs), a family of transmembrane enzymes that modify an estimated 10% of the proteome. At this point in time, no structure of a protein in this family has been solved, and therefore there is poor understanding about the regulation of the enzymes and their substrates. Most proteins, including palmitoylation enzymes and substrates, have …

Engineering Small Molecule Based Dimerization To Induce Translation And Provide Optogenetic Control, Catherine Wright

Chemistry and Chemical Biology ETDs

We wanted to develop a system that combines the spatial control of photoactivation and control of translation to build a tool to spatially control translation in neurons. This kind of tool could be used to investigate the role of spatially controlled translation of any protein in neural behavior. In this way the development and growth of neural processes could be studied to elucidate the mechanisms for spatially sensitive events such as pathfinding, repair, or long-term potentiation.

Chemically induced dimerization was used to install a switch into the activation of translation for specific genes. An abscisic acid (ABA) dependent dimerization of …