Structural Biology Commons^™

Role Of Nucleic Acids As Chaperones In Protein Folding, Zijue Huang

Electronic Theses and Dissertations

Many proteins have slow folding times in vitro that are physiologically untenable. To combat this challenge, ATP-dependent chaperonins are thought to possess the unique ability to catalyze protein folding. Performing quantitative model selection using protein folding and unfolding data, we here show that short nucleic acids containing Gquadruplex (G4) structure can also catalyze protein folding. Performing the experiments as a function of temperature demonstrates that the G4 reshapes the underlying driving forces of protein folding. To understand the structural basis of this catalytic activity, we introduce NMR method to solve the structures, at base-level resolution, of a multiconformer G4 with …

Unraveling The Role Of Hid-1: Insights Into Sgii Regulation, Breanna L. Sellers

Electronic Theses and Dissertations

The regulated secretory pathway is important for the secretion of hormones and neuropeptides from large dense core vesicles (LDCVs). Dysregulation of the biogenesis of LDCVs can result in metabolic and neurological diseases. Recent studies have suggested that the peripheral membrane protein HID-1 plays a role in the sorting and storage of granins which contribute to LDCV biogenesis. To further understand how HID-1 influences the regulation of granins, we investigated potential HID-1 interactors. We determined that while HID-1 is likely functioning in the same pathway as USP25, a deubiquitinating enzyme (DUB), the cause of reduced cellular SgII levels is likely not …

Omani Camels From A Cultural And Genomics Perspective, Al Muatasim Al Zadjali

Electronic Theses and Dissertations

The Dromedarian camel, Camelus dromedarius, is native to the Arabian Peninsula, including the Sultanate of Oman. These camels are used for food, milk, as well as show and racing competitions. Despite their economic and cultural importance research on camels in Oman is limited. The goal of this study was to examine their genomic variation, relationship with camels in other parts of the Arabian Peninsula, and to determine if selective breeding has led to the establishment of distinct breeds in Oman. Information was compiled from multiple sources to produce a comprehensive review on the breeding, management, economic and cultural use, …

Flavonol Glucosylation: A Structural Investigation Of The Flavonol Specific 3-O Glucosyltransferase Cp3gt, Aaron S. Birchfield

Electronic Theses and Dissertations

Flavonoid glycosyltransferases (GTs), enzymes integral to plant ecological responses and human pharmacology, necessitate rigorous structural elucidation to decipher their mechanistic function and substrate specificity, particularly given their role in the biotransformation of diverse pharmacological agents and natural products. This investigation delved into a comprehensive exploration of the flavonol 3-O GT from Citrus paradisi (Cp3GT), scrutinizing the impact of a c-terminal c-myc/6x histidine tag on its enzymatic activity and substrate specificity, and successfully achieving its purification to apparent homogeneity. This established a strong foundation for potential future crystallographic and other structure/function analyses. Through the strategic implementation of site-directed mutagenesis, a thrombin …

Spatial Structure Formation By The Post-Transcriptional Gene Regulator Rsme In Pseudomonas Fluorescens Pf0-1, Anton Evans

Electronic Theses and Dissertations

Microorganisms are often found in microbial communities we call biofilms. Organisms living in these crowded environments have significant evolutionary pressure to retain access to the resources necessary to sustain life. My research uses the bacterium Pseudomonas fluorescens Pf0-1 to study how organisms evolve strategies to solve this crowding problem as aging colonies repeatedly generate mutant patches. These mutants expand the reach of the colony resulting in decreased local density as they push themselves up to the resource rich surface. These spatial structures result from social interactions between the mutant and the parental cells mediated through extracellular secretions, resulting in the …

Applications Of Nuclear Magnetic Resonance Spectroscopy: From Drug Discovery To Protein Structure And Dynamics., Mark Vincent C. Dela Cerna

Electronic Theses and Dissertations

The versatility of nuclear magnetic resonance (NMR) spectroscopy is apparent when presented with diverse applications to which it can contribute. Here, NMR is used i) as a screening/ validation tool for a drug discovery program targeting the Phosphatase of Regenerating Liver 3 (PRL3), ii) to characterize the conformational heterogeneity of p53 regulator, Murine Double Minute X (MDMX), and iii) to characterize the solution dynamics of guanosine monophosphate kinase (GMPK). Mounting evidence suggesting roles for PRL3 in oncogenesis and metastasis has catapulted it into prominence as a cancer drug target. Yet, despite significant efforts, there are no PRL3 small molecule inhibitors …

Molecular Dynamics Simulations Provide Insight Into Stability Of Hyperthermophilic Endoglucanases, Logan E. Sheffield

Electronic Theses and Dissertations

Endoglucanases play a key role in the industrial production of bioethanol, but the most efficient method requires the utilization of high temperatures and is currently limited by the thermostability of endoglucanases. For this reason, it would be beneficial to discover more high-efficiency, thermostable enzymes to utilize in the hydrolytic process. In this study molecular dynamics simulations were performed on structurally similar endoglucanases with varying levels of thermostability to gain insight on what factors contribute to thermostability in endoglucanases. RMSD, RMSF, PCA, hydrogen bonding and salt bridges were analyzed. Finally, protein energy networks were constructed from nonbonded interaction potentials and analysis …

Rna Structure And Function: Biological Relevance In Neurodegenerative And Infectious Disease Pathogenesis, Joshua Imperatore

Electronic Theses and Dissertations

The studies outlined in this dissertation encompass a broad focus, relating to the pathogenesis of neurological disorders such as Alzheimer’s disease, fragile X syndrome, and amyotrophic lateral sclerosis, as well as a novel infectious disease, COVID-19. However, all four studies detailed within this dissertation contain similar elements, namely, the coordinated role of RNA structure and function in essential molecular mechanisms. The results of these investigations allow us to speculate on various disease- related mechanisms, including dysregulated microRNA processing pathways, nucleocytoplasmic shuttling via distinct RNA-protein interactions, and fine-tuned molecular switches controlling genomic dimerization and viral hijacking of host cellular microRNAs. Taken …

Transcription Regulation Of Human Il1b Gene In Monocytes And Lymphoid Cd4 T Cells, Sree H. Pulugulla

Electronic Theses and Dissertations

Cytokines are key regulators of the inflammatory response and play an important role in facilitating intercellular communication between various immune cell types. Interleukin‑1β (IL‑1β) is a potent pro-inflammatory cytokine that is required for robust initiation of innate immune response and subsequent development of adaptive immunity. IL-1β is first synthesized as an inactive cytoplasmic, non‑glycosylated, precursor molecule (proIL‑1β) by monocytes and macrophages in response to invading pathogenic microbes. The activation of caspase‑1 by inflammasomes cleaves proIL-1β into mature biologically active IL-1β that is released from cells via a non-classical, endoplasmic reticulum‑independent secretory pathway directly from the cytoplasm via Gasdermin D membrane …

The N-Terminal Methyltransferase Homologs Nrmt1 And Nrmt2 Exhibit Novel Regulation Of Activity Through Heterotrimer Formation., Jon David Faughn

Electronic Theses and Dissertations

Protein, DNA, and RNA methyltransferases have an ever-expanding list of novel substrates and catalytic activities. Even within families and between homologs, it is becoming clear the intricacies of methyltransferase specificity and regulation are far more diverse than originally thought. In addition to specific substrates and distinct methylation levels, methyltransferase activity can be altered through formation of complexes with close homologs. This work involves the N-terminal methyltransferase homologs NRMT1 and NRMT2. NRMT1 is a ubiquitously expressed distributive trimethylase. NRMT2 is a monomethylase expressed at low levels and in a tissue-specific manner. They are both nuclear methyltransferases with overlapping target consensus sequences …

Quantitative Studies Of Amyloidogenic Protein Residue Interaction Networks And Abnormal Ammonia Metabolism In Neurotoxicity And Disease, Jeddidiah Griffin

Electronic Theses and Dissertations

Investigating similarities among neurological diseases can provide insight into disease processes. Two prominent commonalities of neurological diseases are the formation of amyloid deposits and altered ammonia and glutamate metabolism. Computational techniques were used to explore these processes in several neurological diseases. Residue interaction networks (RINs) abstract protein structure into a series of nodes (representing residues) and edges (representing connections between residues likely to interact). Analyzing the RINs of monomeric forms of amyloidogenic proteins for common network features revealed similarities not previously known. First, amyloidogenic variants of lysozyme were used to demonstrate the usefulness of RINs to the study of amyloidogenic …

Type Ix Secretion System: Characterization Of An Effector Protein And An Insight Into The Role Of C-Terminal Domain Dimeration In Outer Membrane Translocation., Lahari Koneru

Electronic Theses and Dissertations

Porphyromonas gingivalis and Tannerella forsythia are two of the primary pathogens that are associated in the etiology and progression of chronic periodontitis. In T. forsythia, KLIKK proteases are the recently identified group of proteolytic enzymes that are secreted through Type IX secretion system (T9SS). Among, these KLIKK proteases a synergistic relationship was observed between karilysin and mirolysin in invading the host complement system for the survival of the bacteria. Since, karilysin has been already characterized, in this study we propose to study about mirolysin through structural, biochemical and biological characterization. The obtained results from the experiments has shown the …

Neuronal Gq Structures In Neurodegeneration, Damian S. Mcaninch

Electronic Theses and Dissertations

This study investigates protein nucleic acid interactions between various proteins and G quadruplex (GQ) forming messenger RNAs (mRNAs) in human neurological disorders. GQ structures are formed in DNA/RNA, when four guanine residues form planar tetrads stabilized by Hoogsteen base pairing, that stack forming a GQ structure stabilized by potassium ions. These GQ structures are targeted by the arginine-glycine-glycine (RGG) repeat domain containing RNA-binding domain.

Three RGG domain containing RNA-binding proteins, all of which have been implicated in neurological disorders, and their interactions with GQ forming mRNAs, were investigated in this study: fused in sarcoma (FUS), fragile X mental retardation protein …

Bioengineering The Expression Of Active Recombinant Human Cathepsin G, Enteropeptidase, Neutrophil Elastase, And C-Reactive Protein In Yeast, Eliot T. Smith

Electronic Theses and Dissertations

The yeasts Pichia pastoris and Kluyveromyces lactis were used to express several recombinant human proteins for further biochemical characterization. Two substitution variants of recombinant human enteropeptidase light chain (rhEPL) were engineered to modify the extended substrate specificity of this serine protease. Both were secreted as active enzymes in excess of 1.7 mg/L in P. pastoris fermentation broth. The substitution variant rhEPL R96Q showed significantly reduced specificities for the preferred substrate sequences DDDDK and DDDDR; however, the rhEPL Y174R variant displayed improved specificities for these substrate sequences relative to all other reported variants of this enzyme. The neutrophil serine proteases human …

Assessing Crosstalk Between Calcium And Camp In Pancreatic Alpha Cells, Julianne Michelle Imseis

Electronic Theses and Dissertations

Blood glucose levels are regulated by the interactions between insulin and glucagon. Pancreatic alpha cells release glucagon in response to a drop in blood sugar. Pancreatic alpha cells appear to be regulated through multiple signaling pathways. One potential method of regulation occurs from α-MSH or epinephrine stimulated production of the intracellular messenger cyclic AMP. Exocytosis of dense core vesicles containing glucagon ultimately requires an increase in cytosolic calcium. This study focused on the link between cyclic AMP and calcium in pancreatic alpha cells which allows alpha cell regulation. Interactions between calcium and cyclic AMP in pancreatic alpha cells consists of …