Engineering Commons^™

Characterization Of Lignin Structural Variability And The Associated Application In Genome Wide Association Studies, Nathan D. Bryant

Doctoral Dissertations

Poplar (Populus sp.) is a promising biofuel feedstock due to advantageous features such as fast growth, the ability to grow on marginal land, and relatively low lignin content. However, there is tremendous variability associated with the composition of biomass. Understanding this variability, especially in lignin, is crucial to developing and implementing financially viable, integrated biorefineries. Although lignin is typically described as being comprised of three primary monolignols (syringyl, guaiacyl, p-hydroxyphenyl), it is a highly irregular biopolymer that can incorporate non-canonical monolignols. It is also connected by a variety of interunit linkages, adding to its complexity. Secondary cell wall …

Understanding Lignin’S Fast Pyrolysis Through Examination Of The Thermolysis Mechanisms Of Model Oligomers, Ross Wesley Houston

Doctoral Dissertations

The lignocellulosic biorefinery is a visionary concept that endeavors to provide an alternative to fossil-based refineries by producing biobased fungible fuels and specialty chemicals almost exclusively derived currently from petroleum refineries. This vision of the lignocellulosic biorefinery can only be realized if all fractions of lignocellulosic biomass are efficiently deconstructed and valorized to generate a diverse portfolio of products to sustain it against market vicissitudes. Of the three main structural constituents of lignocellulosic biomass (i.e., cellulose, hemicellulose, and lignin), lignin is underutilized despite being the most abundant renewable source of aromatic platform chemicals, representing a growing 250 billion dollar market. …

A Rapid And Ultra-Sensitive Biosensing Platform Based On Tunable Dielectrophoresis For Robust Poc Applications, Yu Jiang

Doctoral Dissertations

With the ongoing pandemic, there have been increasing concerns recently regarding major public health issues such as abuse of organophosphorus compounds, pathogenic bacterial infections, and biosecurity in agricultural production. Biosensors have long been considered a kernel technology for next-generation diagnostic solutions to improve food safety and public health. Significant amounts of effort have been devoted to inventing novel sensing mechanisms, modifying their designs, improving their performance, and extending their application scopes. However, the reliability and selectivity of most biosensors still have much to be desired, which holds back the development and commercialization of biosensors, especially for on-site and point-of-care (POC) …

Elucidating Mammalian Cellular Responses To The Uptake Of Nanoparticles (Nps), Pathogens, And Lipoproteins: Similarities And Differences, Monireh Asoudeh

Doctoral Dissertations

Soft poly-ethylene-glycol (PEG)-based soft nanoparticles (NPs) including cylindrical (CNPs) micelles, spherical (SNPs) micelles, and lipid bilayer vesicles (LNPs) are thought to be treated as foreign objects by mammalian phagocytes. If this hypothesis is true, NPs should trigger a proinflammatory, autophagic phenotype that is similar to the one seen when macrophages phagocytose pathogens or when macrophage surface expressed proteins bind pathogen surface factors such as lipopolysaccharide (LPS). Here, we show that macrophage responses to the above NPs are almost completely unique from those triggered by group A streptococcus (GAS) pathogens (JRS4 cells) and LPS. Instead, macrophages treat these soft NPs more …

Crowd Control: Regulating The Spatial Organization Of Biopolymers And Gene Expression By Macromolecular Crowding, Gaurav Chauhan

Doctoral Dissertations

The intracellular environment is crowded with macromolecules that can occupy a significant fraction of the cellular volume. This can give rise to attractive depletion interactions that impact the conformations and interactions of biopolymers, as well as their interactions with confining surfaces. We used computer simulations to study the effects of crowding on biologically-inspired models of polymers. We showed that crowding can lead to attractive interactions between two flexible ring polymers, and we further characterized the adsorption of both flexible and semiflexible polymers onto confining surfaces. These results indicate that crowding-induced depletion interactions could play a role in the spatial organization …

Interactions Between Soft Nanoparticles And Mammalian Cells, Mitchell Raith

Doctoral Dissertations

Nanoparticles have been of interest to the pharmaceutical industry since the 1980s. The first FDA approved nanoparticle-based therapies included liposomal anesthesia agents. Since then, the amount of FDA-approved nanoparticle therapies remains low. This is because nanoparticle-patient interactions can be very complex and are not well understood. Complicating factors also include increasing obesity rates among the patient population and many small animal pre-clinical trials are completed with healthy, lean animals. The biochemical differences between lean and obese patients prevents early studies from accurately predicting nanoparticle clinical behaviors. Many nanoparticles fail in trails. In this thesis, I aimed to uncover how nanoparticles …

Improving The Biocompatibility Of The Bio-Inorganic Interface For Enhanced Photosystem I-Based Biophotovoltaic Device Performance, Alexandra H. Teodor

Doctoral Dissertations

The world’s energy demands are projected to increase by nearly 50% by the year 2040, and consumption of carbon-based fuels continues to release greenhouse gases such as carbon dioxide and methane into the atmosphere. This has been causally linked with climate change and increased extreme weather events, which has been further linked to adverse health outcomes and negative effects on biodiversity, food security, and increased disease transmission. Clearly, there is a need for a sustainable, carbon-free, and cost-effective method of energy production to meet growing energy production demands. The sun irradiates Earth’s surface annually with ~80,000 terawatts (TW), making solar …

Elucidating Mechanisms And Genotypes Underlying Robust Phenotypes In Yarrowia Lipolytica, Caleb M. Walker

Doctoral Dissertations

Robustness is an important phenotype for bioenergy microbes to acquire but is difficult to engineer. Hence, tools for engineering microbial robustness are critical to unlock novel phenotypes for innovative bioprocessing strategies. The oleaginous yeast, Yarrowia lipolytica, is an exceptionally robust microbe that can tolerate stressful environments, assimilate a wide range of substrates, and produce high-value chemicals. In this doctoral dissertation, the impacts of systems biology and metabolic engineering to reveal mechanisms and identify genotypes- underlying robust phenotypes are addressed.

The first approach employs adaptive laboratory engineering to generate a platform strain by which to study superior robust mechanisms. This …

Engineering Modularity Of Ester Biosynthesis Across Biological Scales, Hyeongmin Seo

Doctoral Dissertations

Metabolic engineering and synthetic biology enable controlled manipulation of whole-cell biocatalysts to produce valuable chemicals from renewable feedstocks in a rapid and efficient manner, helping reduce our reliance on the conventional petroleum-based chemical synthesis. However, strain engineering process is costly and time-consuming that developing economically competitive bioprocess at industrial scale is still challenging. To accelerate the strain engineering process, modular cell engineering has been proposed as an innovative approach that harnesses modularity of metabolism for designing microbial cell factories. It is important to understand biological modularity and to develop design principles for effective implementation of modular cell engineering. In this …

Engineered Switch Protein Inspired By Novel Protein Affinity Transition Mode, Liang Fang

Doctoral Dissertations

Many natural proteins involved in complex biological processes such as ligand binding and protein folding demonstrate multiple, allosterically-regulated conformational states, with protein activity regulated by effector molecules. The alpha L integrin and its inserted domain (I domain) is one example of such a protein. The binding of the effector molecule such as talin or filamin to the cytoplasmic domain of the integrin increases the binding affinity between I- domain and its ligand intercellular adhesion molecule-1, known as ICAM-1.There are multiple models attempting to describe the mechanism responsible for the change in binding affinity. According to research conducted by our lab, …

Rapid Design, Construction, And Validation Of Synthetic Metabolic Pathways In A Modular Escherichia Coli (Chassis) Cell, Jong-Won Lee

Doctoral Dissertations

Current strain development has been hindered by the vast biochemical space in nature. The concept of modular cell design has been invented to enable rapid and predictable construction of multiple optimal production strains for efficient production of a large variety of biochemicals with minimal experimental effort. While modular cell design principles have been successfully validated in some cases, its development is still limited by the small library of the production modules demonstrated.

The goals of this thesis are i) to establish a framework for rapid design, construction, and validation of production modules to explore a large space of molecules (e.g., …

Approaches To Studying Bacterial Biofilms In The Bioeconomy With Nanofabrication Techniques And Engineered Platforms., Michelle Caroline Halsted

Doctoral Dissertations

Studies that estimate more than 90% of bacteria subsist in a biofilm state to survive environmental stressors. These biofilms persist on man-made and natural surfaces, and examples of the rich biofilm diversity extends from the roots of bioenergy crops to electroactive biofilms in bioelectrochemical reactors. Efforts to optimize microbial systems in the bioeconomy will benefit from an improved fundamental understanding of bacterial biofilms. An understanding of these microbial systems shows promise to increase crop yields with precision agriculture (e.g. biosynthetic fertilizer, microbial pesticides, and soil remediation) and increase commodity production yields in bioreactors. Yet conventional laboratory methods investigate these micron-scale …

Morphological And Photoelectrochemical Characterization Of Membrane Reconstituted Photosystem I (Psi), Seyedeh Hanieh Niroomand

Doctoral Dissertations

The robust structural and photoactive electrochemical properties of Photosystem I (PSI), a transmembrane photosynthetic protein complex, make it an ideal candidate for incorporation into solid state bioelectronic or hybrid photovoltaic devices. However, the first step towards the successful fabrication of such devices requires systematic assembly of oriented and functional PSI onto desired bio-abio interfaces via suitable protein scaffoldings. Hence, this dissertation focuses on utilizing the cyanobacterial PSI for integration into organic/inorganic interfaces that mediate photo-electrochemical energy conversions for electricity and/or solar fuel production. To this end, in this study the effect of systematic incorporation of PSI complexes into synthetic membrane-bound …

Elucidating The Effects Of Metabolic State On Nanoparticle Distribution In Mice And In Vitro Uptake, Kevin James Quigley

Doctoral Dissertations

Since almost 70% of the U.S. population over 20 years old is overweight and 30% is obese, with much of the western world following suit, many patients that will potentially be administered circulating nanoparticles designed to localize to tumors and avoid non-target areas will have significant amounts of white adipose tissue (WAT), enlarged livers, and additional metabolic complications such as type 2 diabetes. However, studies on nanoparticle biodistribution and efficacy take place almost without exception in lean rodents with healthy metabolic states. In this work, I determined the biodistribution of model nanoparticles – neutral filomicelles and polystyrene spheres both carrying …

In Silico Driven Metabolic Engineering Towards Enhancing Biofuel And Biochemical Production, Richard Adam Thompson

Doctoral Dissertations

The development of a secure and sustainable energy economy is likely to require the production of fuels and commodity chemicals in a renewable manner. There has been renewed interest in biological commodity chemical production recently, in particular focusing on non-edible feedstocks. The fields of metabolic engineering and synthetic biology have arisen in the past 20 years to address the challenge of chemical production from biological feedstocks. Metabolic modeling is a powerful tool for studying the metabolism of an organism and predicting the effects of metabolic engineering strategies. Various techniques have been developed for modeling cellular metabolism, with the underlying principle …

Elucidating Mechanisms Of Lipid Droplet Formation In The Fission Yeast, Schizosaccharomyces Pombe, Alexander William Meyers

Doctoral Dissertations

Cellular function relies on the proper sequestration of fats in organelles called lipid droplets. Lipid droplet metabolism is inherently linked to many disorders including obesity, type-2 diabetes, and atherosclerosis, so further elucidation of the bio-physical phenomena governing these diseases, is crucial for their respective treatments.

Once widely regarded as inert, these neutral lipid storage depots are highly dynamic and are increasingly shown to affect a wide array of biological processes. Droplet formation requires the accumulation of neutral lipids and related factors at specific cellular domains, however because this occurs at nanometer length-scales, details are lacking. Here, we try to provide …

Photosystem I-Based Applications For The Photo-Catalyzed Production Of Hydrogen And Electricity, Rosemary Khuu Le

Doctoral Dissertations

The aim of this dissertation was to optimize systems integrating the photosystem I (PSI) redox protein, which is involved in photosynthesis, with noble metals for electron transfer to show its versatility: 1) in solution coupled with platinum to mediate hydrogen evolution and 2) on a planar gold surface for electricity production.

Response surface methodology was utilized to study variables that affect hydrogen (H₂) yield from platinized-PSI. Light intensity, temperature, and platinum concentration were varied during the platinum-photo-reduction process. Analysis of the effects of the variables on H₂ yield allowed for determination of a condition for optimized hydrogen …

The Structure And Function Of Photosystem I And Photosystem I – Hydrogenase Protein Fusions: An Experimental And Computational Study, Bradley Jordan Harris

Doctoral Dissertations

Photosystem I (PSI) is a membrane protein involved in the photosynthetic cycle of plants, algae, and cyanobacteria that is of specific interest due to its ability to harness solar energy to generate reducing power. This work seeks to form an in vitro hybrid protein fusion between the membrane integral PSI protein and the membrane-bound hydrogenase (MBH) enzyme, in an effort to improve electron transport between these two proteins.

Small-angle neutron scattering (SANS) was used to characterize the detergent-solubilized solution structure of trimeric PSI from the cyanobacterium Thermosynechococcus elongatus, which showed that the detergent interacts primarily with the hydrophobic periphery …

A Tale Of Two Protein Switches: Engineering, Characterizing, And Understanding A Novel And A Natural Switch, James Vincent Price

Doctoral Dissertations

Throughout nature, many proteins provide a specific function in response to some input signal (e.g., phosyphorylation, pH, etc.), a process that is oftentimes described as switching a protein “on” or “off.” The advent of protein engineering has allowed for the creation and understanding of chimeric proteins for uses in a number of applications such as therapeutics, biosensors and energy production. Two proteins, hemagglutinin (HA) of fowl plague influenza virus and a chimeric protein comprising a fusion between the LFA-1 I domain and the EF3 and EF4 hands of calmodulin, have been investigated in this dissertation. Both of these proteins undergo …

Industrial Robustness: Understanding The Mechanism Of Tolerance For The Populus Hydrolysate Tolerant Strain Of Clostridium Thermocellum, Jessica Leigh Linville

Doctoral Dissertations

An industrially robust microorganism that can efficiently degrade and convert lignocellulosic biomass into ethanol and next-generation fuels is required to economically produce future sustainable liquid transportation fuels. The anaerobic, thermophilic, cellulolytic bacterium Clostridium thermocellum is a candidate microorganism for such conversions but it, like many bacteria, is sensitive to potential toxic inhibitors developed in the hydrolysate produced during biomass processing. Microbial processes leading to tolerance of the inhibitory compounds found in the pretreated biomass hydrolysate are likely complex and involve multiple genes. In this study, a 17.5% v/v Populus hydrolysate tolerant mutant strain of C. thermocellum was developed by directed …

Modeling Chemical Degradation And Proton Transport In Perfluorosulfonic Acid Ionomers, Milan Kumar

Doctoral Dissertations

The ionomer-membrane interface in a membrane electrode assembly connects the catalyst and membrane and allows hydrated protons to move between the catalyst and membrane. The continuous operation of the polymer membrane electrolyte fuel cell at high temperature and/or in frequent freeze/thaw cycles leads to membrane degradation and delamination of the interface, which lower the proton conductivity. In this dissertation, we modeled the chemical degradation and proton conductivity of perfluorosulfonic acid (PFSA) ionomers by ab initio calculations and macroscopic modeling. All ab initio calculations were performed using Gaussian 03 suites of program by employing B3LYP/6-311++G** method/basis set. The macroscopic modeling involves …

Bioreactor System Designs For Lipase-Catalyzed Synthesis Of Saccharide- Fatty Acid Esters In Solvent-Free Media, Ran Ye

Doctoral Dissertations

As nontoxic biobased surfactants derived from plant oils and cellulose or starch, saccharide-fatty acid esters are widely used in cosmetics, food, and pharmaceutical industries due to their biocompatibility, biodegradability as well as antimicrobial activity. Generally, saccharide-fatty acid esters are synthesized chemically under high pressure, temperature and the presence of alkaline or acid catalysts leading to low-quality products (chemo-degradation of double bonds and oxygenated moieties) and large amounts of byproducts. In contrast, biocatalytic synthesis enhances sustainability: near-ambient pressure and temperature, the absence of toxic, acids and bases catalysts, and improved selectivity of products. For lipase-catalyzed synthesis under nearly anhydrous conditions, the …

Protein Engineering For The Enhanced Photo-Production Of Hydrogen By Cyanobacterial Photosystem I, Ifeyinwa Jane Iwuchukwu

Doctoral Dissertations

Photosystem I (PSI) from plants, algae, and cyanobacteria can mediate H2 evolution in vivo and in vitro. A simple, self-platinization procedure that permits stable PSI-mediated H2 evolution in vitro has been developed. The H2 evolution capabilities of PSI from Thermosynechococcus elongatus have been characterized. This organism utilizes cytochrome c6 (cyt c6) as the e- donor to P700. Using a solution-based, self-organized platinization of the PSI nanoparticles, this study demonstrates a sodium ascorbate-cyt-PSI-Pt-H2 electron transport and proton reduction system that yields light-dependent H2. The system was thermostable with H2 evolution increasing up to 55°C. In addition, stability studies have shown the …

Molecular Simulations Of Adsorption And Diffusion In Metal-Organic Frameworks (Mofs), Ruichang Xiong

Doctoral Dissertations

Metal-organic frameworks (MOFs) are a new class of nanoporous materials that have received great interest since they were first synthesized in the late 1990s. Practical applications of MOFs are continuously being discovered as a better understanding of the properties of materials adsorbed within the nanopores of MOFs emerges. One such potential application is as a component of an explosive-sensing system. Another potential application is for hydrogen storage.

This work is focused on tailoring MOFs to adsorb/desorb the explosive, RDX. Classical grand canonical Monte Carlo (GCMC) and molecular dynamic (MD) simulations have been performed to calculate adsorption isotherms and self-diffusivities of …

Isolation And Thermodynamic Characterization Of Aminoglycoside Nucleotidyltransferase (2″)-Ia, Roger Edward Wright

Doctoral Dissertations

Aminoglycoside nucleotidyltransferase(2″)-Ia is one of the most important aminoglycoside-modifying enzymes. Because of the difficulties in isolating this enzyme with a high level of purity, very little work has been reported for this enzyme. A procedure for obtaining this enzyme with a purity level of greater than 95% from inclusion bodies was developed. The optimal conditions for isolating ANT(2″) were solubilizing the inclusion bodies in 8 M urea followed by direct dilution of the enzyme into 0.1 M Tris-HCl pH 8.5, 0.2 M KCl, 0.4 ML-arginine and 5 mM reduced glutathione at 4°C. The determination of an effective method of obtaining …