Theses and Dissertations from UMD
Permanent URI for this communityhttp://hdl.handle.net/1903/2
New submissions to the thesis/dissertation collections are added automatically as they are received from the Graduate School. Currently, the Graduate School deposits all theses and dissertations from a given semester after the official graduation date. This means that there may be up to a 4 month delay in the appearance of a give thesis/dissertation in DRUM
More information is available at Theses and Dissertations at University of Maryland Libraries.
Browse
6 results
Search Results
Item 3D ENGINEERING OF VIRUS-BASED PROTEIN NANOTUBES AND RODS: A TOOLKIT FOR GENERATING NOVEL NANOSTRUCTURED MATERIALS(2018) Brown, Adam Degen; Culver, James N; Bioengineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Technological innovation at the nanometer scale has the potential to improve a wide range of applications, including energy storage, sensing of environmental and medical signals, and targeted drug delivery. A key challenge in this area is the ability to create complex structures at the nanometer scale. Difficulties in meeting this challenge using traditional fabrication methods have prompted interest in biological processes, which provide inspiration for complex structural organization at nanometer to micrometer length scales from self-assembling components produced inexpensively from common materials. From that perspective, a system of targeted modifications to the primary amino acid structure of Tobacco mosaic virus (TMV) capsid protein (CP) has been developed that induces new self-assembling behaviors to produce nanometer-scale particles with novel architectures. TMV CPs contain several negatively charged carboxylate residues which interact repulsively with those of adjacent CP subunits to destabilize the assembled TMV particle. Here, the replacement of these negatively charged carboxylate residues with neutrally charged or positively charged residues results in the spontaneous assembly of bacterially expressed CP into TMV virus-like particles (VLPs) with a range of environmental stabilities and morphologies and which can be engineered to attach perpendicularly to surfaces and to display functional molecular patterns such as target-binding peptide chains or chemical groups for attachment of functional targets. In addition, the distinct electrostatic surface charges of these CP variants enable the higher-level coassembly of TMV and VLP into continuous rod-shaped nanoparticles with longitudinally segregated distribution of functionalities and surface properties. Furthermore, the unique, novel, environmentally responsive assembly and disassembly behaviors of the modified CPs are shown to act as simple mechanisms to control the fabrication of these hierarchically structured functional nanoparticles.Item Ribosomal Protein L11: A Cog in the Nanomachine(2011) Rhodin, Michael Hoover Johannes; Dinman, Jonathan D; Molecular and Cell Biology; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Comprised of two major subunits of both rRNA and proteins, the ribosome is a biological nanomachine, acting as the central player in the process of protein translation. Recent advances in molecular imaging have enabled the visualization of the disparate functional centers within the ribosome, leading to the question of how these critical regions coordinate their actions and communicate with each other. This work examines the essential ribosomal protein L11, located in the central protuberance of the large subunit. L11 maintains connections with the 5S rRNA, H84 of the 25S rRNA, comes in close proximity to the T-loop of the bound peptidyl tRNA, and shares an intersubunit bridge with small subunit protein S18. L11 was found to have a critical dynamic loop which samples the occupancy status of the P-site pocket of the ribosome and communicates this information through H84. L11's intersubunit bridge (the B1b/c bridge) mediates an intersubunit communication network from the decoding center to the peptidyl transferase center of the ribosome. L11 is also involved in proper subunit joining. Mutations in L11 were found to have effects on A- and P-site tRNA binding, translational fidelity, and growth and viability of yeast cells.Item Host Molecular Responses in Chickens Infected with an Avian Influenza Virus(2008-11-20) Ramirez-Nieto, Gloria Consuelo; Perez, Daniel R.; Veterinary Medical Science; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Avian influenza virus has a segmented RNA genome that allows the virus to evolve continuously and generate new strains. Wild birds serve as natural reservoirs of avian influenza virus and provide a potential source for emergence of new viruses, which traverse host barriers and infect new avian or mammalian species. The mechanisms involved in this process are not completely understood. Our main goal is to understand host-pathogen interactions involved in avian influenza pathogenicity. As part of our approach we studied the effect of pre-exposure of chickens to IBDV (infectious bursal disease virus) on host susceptibility to infection, disease progression, and host molecular responses to infection with a mallard H5N2 low pathogenic avian influenza (LPAI) virus. We found that prior exposure of chickens to IBDV led to increased susceptibility to infection with the mallard H5N2 LPAI virus compared to normal chickens. This increased susceptibility allowed us to further adapt the virus to chickens. After 22 passages (P22) in IBDV-pre-exposed chickens, the LPAI virus replicated substantially better than the wild-type (WT) mallard virus in both IBDV-exposed and normal chickens. Interestingly, the P22 virus showed similar levels of replication in the respiratory and intestinal tracts of both groups, although it caused exacerbated signs of disease and severe lesions in the IBDV-pre-exposed group. We suggest that prior IBDV exposure provides a port of entry for avian influenza in an otherwise resistant chicken population. Furthermore, adaptation of avian influenza (AI) in IBDV-exposed chickens may allow for the selection of AI virus strains with expanded tissue tropism. We also studied the effects of host response to H5N2 AI in normal and IBDV-infected birds using high-throughput gene expression analysis. We demonstrated that IBDV-exposed chickens showed less than optimal humoral responses to LPAI infection as well as alterations in local molecular pathways that eventually led to exacerbated disease and death. At the molecular level we found amino acid substitutions in the surface glycoprotein hemagglutinin (HA). Those changes suggest selection for a virus that binds to and replicates more efficiently in chickens. Taken together our results suggest that IBDV-pre-exposure may play a role in exacerbating AI-induced pathogenicity.Item Programmed Ribosomal Frameshifting in SARS-CoV and HIV-1(2007-12-10) Neeriemer, Jessica; Dinman, Jonathan D; Cell Biology & Molecular Genetics; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Programmed ribosomal frameshifting controls the ratio of two protein products made in a variety of viruses and mammalian cells. This occurs when the ribosome is translating mRNA, pauses at secondary structure, slips back one base in the 5' direction, and continues translation in a new reading frame. A series of SARS-CoV pseudoknot mutants were generated to examine important features of frameshifting, and an antibiotic was tested for its effect on HIV and SARS-CoV frameshifting. Other mutants were made in the human CCR5 gene to determine whether frameshifting occurs. It was found that mRNA stability and unpaired adenosines influence frameshifting, and increasing concentrations of the antibiotic gentamicin increases frameshifting. Moreover, CCR5, the co-receptor for HIV, contains a working frameshifting signal. This study pinpoints several antiviral targets and important factors for HIV and SARS-CoV pathogenesis.Item Nanopatterning of Recombinant Proteins and Viruses Using Block Copolymer Templates(2007-01-19) Cresce, Arthur von Wald; Kofinas, Peter; Material Science and Engineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)The study of interfaces is important in understanding biological interactions, including cellular signaling and virus infection. This thesis is an original effort to examine the interaction between a block copolymer and both a protein and a virus. Block copolymers intrinsically form nanometer-scale structures over large areas without expensive processing, making them ideal for the synthesis of the nanopatterned surfaces used in this study. The geometry of these nanostructures can be easily tuned for different applications by altering the block ratio and composition of the block copolymer. Block copolymers can be used for controlled uptake of metal ions, where one block selectively binds metal ions while the other does not. 5-norbornene-2,3-dicarboxylic acid is synthesized through ringopening metathesis polymerization. It formed spherical domains with spheres approximately 30 nm in diameter, and these spheres were then subsequently loaded with nickel ion. This norbornene block copolymer was tested for its ability to bind histidine-tagged green fluorescent protein (hisGFP), and it was found that the nickel-loaded copolymer was able to retain hisGFP through chelation between the histidine tag and the metal-containing portions of the copolymer surface. Poly(styrene-b-4-vinylpyridine) (PS/P4VP) was also loaded with nickel, forming a cylindrical microstructure. The binding of Tobacco mosaic virus and Tobacco necrosis virus was tested through Tween 20 detergent washes. Electron microscopy allowed for observation of both block copolymer nanostructures and virus particles. Results showed that Tween washes could not remove bound Tobacco mosaic virus from the surface of PS/P4VP. It was also seen that The size and tunability of block copolymers and the lack of processing needed to attain different structures makes them attractive for many applications, including microfluidic devices, surfaces to influence cellular signaling and growth, and as a nanopatterning surface for organized adhesion.Item Wiring the ribosome: functions of ribosomal proteins L3 and L10, and 5S rRNA(2006-09-29) Petrov, Alexey; Dinman, Jonathan D; Cell Biology & Molecular Genetics; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)The ribosome is a megadalton complex that performs protein synthesis with tremendous speed and accuracy. Atomic resolution ribosome structures have been resolved within the last five years. These have provided the 3-dimensional locations of all ribosomal components, and have revealed structures of the active centers. However, the precise mechanisms of the various functions performed by the ribosome are still unknown. This work is an attempt to understand some of the functional relationships between different active centers of the ribosome (or the "wiring" of the ribosome), and mechanisms by which such communication occurs. Here we present an analysis of three ribosomal components: ribosomal proteins L3 and L10, and 5S rRNA. Studies of L3 suggest that accommodation of aminoacyl-tRNAs (aa-tRNA) may be the mechanism that induces the "active" conformation of the peptidyl transferase center. We have proposed a mechanism in which rRNA movement associated with aa-tRNA accommodation facilitates conformational changes in the peptidyl transferase center (PTC) through the formation of a network of hydrogen bond interactions. A saturation mutagenesis analysis of 5S rRNA disproves the previous notion that 5S rRNA is a resilient molecule. An analysis of naturally occurring 5S rRNA variants suggests that this molecule may participate in posttranscriptional regulation of gene expression via the nonsense-mediated mRNA decay (NMD) pathway. Lastly, a random mutagenesis analysis of ribosomal protein L10 has resulted in the creation of a powerful toolbox that will be used for elucidation of ribosome export/maturation pathways. Future structure/functional analyses of these mutants may also help to reveal roles of helices 38 and 89 of 25S rRNA.