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Title
Engineering approaches to Shewanella taxis studies
Creator
Li, Rui
Date
2012
Collection
Electronic Theses & Dissertations
Description
Shewanella species are famous for their broad range of terminal electron acceptors and the ability to perform taxis towards both soluble and insoluble electron acceptors, which may help explain their nearly ubiquitous presence in widely disparate environmental niches around the world. Studies of Shewanella's tactic properties are important to understand their competitiveness and roles in elemental (e.g. nitrogen, sulfur, iron, manganese and others) cycling...
Show moreShewanella species are famous for their broad range of terminal electron acceptors and the ability to perform taxis towards both soluble and insoluble electron acceptors, which may help explain their nearly ubiquitous presence in widely disparate environmental niches around the world. Studies of Shewanella's tactic properties are important to understand their competitiveness and roles in elemental (e.g. nitrogen, sulfur, iron, manganese and others) cycling and bioremediation (e.g., precipitation of soluble uranium oxides). Population-level microbial taxis involves a complex interplay of cellular process (e.g., growth, metabolism, chemotaxis, and random motility) and molecular processes (e.g., diffusion of electron donors and acceptors that serve as attractants). This dissertation describes the use of multiple approaches including engineering tools, biological assays and mathematical models to study population-level growth and taxis of Shewanella in response to applied and cell-generated gradients of soluble electron acceptors. The model was able to reproduce key trends of the observed cell growth and migration patterns in either diffusion gradient chamber (DGC) or motility assays, which validate the use of our approaches to measure and simulate Shewanella's taxis in response to electron acceptor gradients.New hypotheses relevant to Shewanella's taxis were investigated to help understand how ecological niches containing various electron acceptor gradients influence the distribution of Shewanella baltica strains with different genotypes and hence different chemotactic behaviors. We studied the impact of opposing gradients of nitrate and fumarate on the chemotactic behaviors of S. oneidensis MR-1 fumarate reductase and nitrate reductase mutants, where the mixture of mutant strains could be partially separated into two populations via formation of two separate chemotactic bands, one moving toward each electron acceptor source. We also studied cell behavior in the presence of insoluble electron acceptor manganese dioxide (MnO2). A novel mechanism, mediated energy taxis, we proposed by which shewanellae use self-secreted riboflavin as both an electron shuttle and an attractant to direct cell movement toward local sources of insoluble electron acceptors. To test this hypothesis, taxis measurements were conducted in the presence of in various insoluble electron acceptors and with chemotaxis mutants. The results strongly supported the hypothesis, and mathematical models based on these mediated energy taxis mechanisms were able to predict experimental trends.
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Title
Properties and applications of self-assembled biomolecules in nanostructured biomimetic interfaces
Creator
Castro Forero, Angelines A.
Date
2011
Collection
Electronic Theses & Dissertations
Description
Living cells are able to synthesize a wide range of biomolecules whose precisely controlled size, shape, charge, linkage, molecular weight, functional groups, and binding affinity enable self-assembly. These molecular building blocks, including amino acids, carbohydrates, lipids, and nucleic acids, form macromolecular structures, whose functional properties often cannot be matched by man-made materials. To harness the advantages of biomolecules in the design of new materials, controlled...
Show moreLiving cells are able to synthesize a wide range of biomolecules whose precisely controlled size, shape, charge, linkage, molecular weight, functional groups, and binding affinity enable self-assembly. These molecular building blocks, including amino acids, carbohydrates, lipids, and nucleic acids, form macromolecular structures, whose functional properties often cannot be matched by man-made materials. To harness the advantages of biomolecules in the design of new materials, controlled methods of fabrication are needed. This dissertation focuses on advancing the understanding of the fabrication process of two biomaterials, phospholipids and self-organizing peptides.Phospholipids, the most abundant components in cell membranes, spontaneously form lipid bilayers in aqueous solution. Synthetic lipid bilayers in the form of small unilamellar vesicles (SUV) and supported bilayer lipid membranes (sBLM) are widely used to study membrane-mediated processes and to mimic natural membranes in surface-based devices. The two preferred methods to fabricate SUVs are extrusion and sonication. This study tested the hypothesis that the SUV fabrication method influenced key bilayer properties. The results confirmed that the morphology, average size and distribution of SUVs varied with the fabrication method. However, the molecular-scale behavior of the bilayer, as studied by fluorescence lifetime, anisotropy measurements, and translational diffusion was independent of the fabrication method.Some peptides self assemble into biologically important macromolecular structures and thus represent excellent candidates for bottom-up fabrication of nanostructured biomimetic materials. This study tested the hypothesis that synthetic analogues of the conductive pili produced by the metal-reducing bacterium Geobacter sulfurreducens could be fabricated in-vitro. Type IV pili are homopolymers of a pilin subunit (PilA) or pilin that polymerizes via hydrophobic interactions to form pili. The resulting synthetic protein nanowires could be potentially used to develop novel nanotechnologies, including nanoelectronic devices. Using genetic engineering and protein expression tools, a method to mass-produce pilA peptides was developed. In-vitro assembly of recombinant pilin subunits resulted in the formation of filaments with properties similar to native G. sulfurreducens pili including a similar diameter, tendency to aggregate into tangled bundles, and electrical conductivity. This achievement offers a potential cost-effective method to mass-produce protein nanowires for commercial applications.Collectively, this research provided important insights into the use of self-assembling biomolecules for bottom-up fabrication of functional and nanostructured biomimetic interfaces. It indicated that the liposome preparation method only affects physical properties of the vesicles (average size and distribution) but not the dynamics of rotational and translational diffusion of the bilayers. It also enabled production of recombinant pilin subunits into synthetic nanowires having properties similar to the native pili of G. sulfurreducens.
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