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- Genre: Masters Thesis
Description
Quiescin sulfhydryl oxidase 1 (QSOX1) is an enzyme that catalyzes disulfide bond formation by oxidizing two free sulfhydryl groups. QSOX1 consists of a thioredoxin (Trx) and an ERV (essential for respiration and viability)/ALR (augmenter of liver regeneration) domain which each contain CxxC motifs that work to bind to substrates and shuttle electrons to a flavin adenine dinucleotide (FAD) cofactor that accepts the electrons and reduces molecular oxygen to hydrogen peroxide. Investigation of the role of QSOX1 in cancer progression started when it was found at higher abundance in pancreatic ductal adenocarcinoma (PDA) patient plasma compared to healthy normal donor plasma. Increased expression in QSOX1 has been further identified in breast, lung, kidney, prostate, and other cancers. QSOX1 expression is associated with cell proliferation and invasion in vitro and tumor growth in vivo. Additionally, the enzymatic activity of QSOX1 in the extracellular matrix (ECM) is important for cell invasion in vitro. Small molecule inhibitors of QSOX1 have been shown to have antitumorigenic properties in vitro and in vivo. It was hypothesized that monoclonal antibodies (mAbs) against QSOX1 would inhibit cell invasion in vitro. In this work, mice were immunized with eukaryotic-derived rQSOX1 for generation of hybridomas. Hundreds of hybridoma clones were screened by enzyme-linked immunosorbent assay (ELISA) and a fluorescent QSOX1 activity assay. Multiple rounds of subcloning and screening identified 2F1.14 and 3A10.6 as mAbs of interest. The genes for the variable regions of the antibodies were rescued and sequenced. The sequences were aligned with the variable region sequences of another published αQSOX1 mAb scFv492.1. 2F1.14 inhibits the enzymatic activity of QSOX1 by binding to the active site of QSOX1, which was determined by epitope mapping against mutants of QSOX1 that contained mutations in the active site. 3A10.6 did not appear to inhibit the function of QSOX1 in the activity assay; however, it, along with 2F1.14, suppressed tumor invasion in a 3D invasion model. These findings support the developing idea that QSOX1 is a viable target for cancer treatment because targeted inhibition of QSOX1 extracellularly reduced invasive activity. The mAbs and rQSOX1 variants produced here can serve as tools in furthering the characterization of QSOX1 and its role in cancer.
ContributorsKoelbel, Calvin John (Author) / Lake, Douglas (Thesis advisor) / Chen, Qiang "Shawn" (Committee member) / Ho, Thai (Committee member) / Arizona State University (Publisher)
Created2019
Description
The public has expressed a growing desire for more sustainable and green technologies to be implemented in society. Bio-cementation is a method of soil improvement that satisfies this demand for sustainable and green technology. Bio-cementation can be performed by using microbes or free enzymes which precipitate carbonate within the treated soil. These methods are referred to as microbial induced carbonate precipitation (MICP) and enzyme induced carbonate precipitation (EICP). The precipitation of carbonate is the formation of crystalline minerals that fill the void spaces within a body of soil.
This thesis investigates the application of EICP in a soil collected from the Arizona State University Polytechnic campus. The surficial soil in the region is known to be a clayey sand. Both EICP and MICP have their limitations in soils consisting of a significant percentage of fines. Fine-grained soils have a greater surface area which requires the precipitation of a greater amount of carbonate to increase the soil’s strength. EICP was chosen due to not requiring any living organisms during the application, having a faster reaction rate and size constraints.
To determine the effectiveness of EICP as a method of improving a soil with a significant amount of fines, multiple comparisons were made: 1) The soil’s strength was analyzed on its own, untreated; 2) The soil was treated with EICP to determine if bio-cementation can strengthen the soil; 3) The soil had sand added to reduce the fines content and was treated with EICP to determine how the fines percentage effects the strength of a soil when treated with EICP.
While the EICP treatment increased the strength of the soil by over 3-fold, the strength was still relatively low when compared to results of other case studies treating sandy soils. More research could be done with triaxial testing due to the samples of the Polytechnic soil’s strength coming from capillarity.
This thesis investigates the application of EICP in a soil collected from the Arizona State University Polytechnic campus. The surficial soil in the region is known to be a clayey sand. Both EICP and MICP have their limitations in soils consisting of a significant percentage of fines. Fine-grained soils have a greater surface area which requires the precipitation of a greater amount of carbonate to increase the soil’s strength. EICP was chosen due to not requiring any living organisms during the application, having a faster reaction rate and size constraints.
To determine the effectiveness of EICP as a method of improving a soil with a significant amount of fines, multiple comparisons were made: 1) The soil’s strength was analyzed on its own, untreated; 2) The soil was treated with EICP to determine if bio-cementation can strengthen the soil; 3) The soil had sand added to reduce the fines content and was treated with EICP to determine how the fines percentage effects the strength of a soil when treated with EICP.
While the EICP treatment increased the strength of the soil by over 3-fold, the strength was still relatively low when compared to results of other case studies treating sandy soils. More research could be done with triaxial testing due to the samples of the Polytechnic soil’s strength coming from capillarity.
ContributorsRoss, Johnathan (Author) / Kavazanjian, Edward (Thesis advisor) / Zapata, Claudia (Committee member) / Hamdan, Nasser (Committee member) / Arizona State University (Publisher)
Created2018
Description
Synthetic biology and metabolic engineering has aided the production of chemicals using renewable resources, thus offering a solution to our dependence on the dwindling petroleum resources. While a major portion of petroleum resources go towards production of fuels, a significant fraction also goes towards production of specialty chemicals. There has been a growing interest in recent years in commercializing bio-based production of such high value compounds. In this thesis the biosynthesis of aromatic esters has been explored, which have typical application as flavor and fragrance additive to food, drinks and cosmetics. Recent progress in pathway engineering has led to the construction of several aromatic alcohol producing pathways, the likes of which can be utilized to engineer aromatic ester biosynthesis by addition of a suitable enzyme from the acyltransferase class. Enzyme selection and screening done in this work has identified chloramphenicol O-acetyltransferase enzyme(CAT) as a potential candidate to complete the biosynthetic pathways for each of 2-phenethyl acetate, benzyl acetate, phenyl acetate and acetyl salicylate. In the end, E. coli strains capable of producing up to 60 mg/L 2-phenethyl acetate directly from glucose were successfully constructed by co-expressing CAT in a previously engineered 2-phenylethanol producing host.
ContributorsMadathil Soman Pillai, Karthika (Author) / Nielsen, David (Thesis advisor) / Wang, Xuan (Committee member) / Torres, Cesar (Committee member) / Arizona State University (Publisher)
Created2016
Description
Universal biology is an important astrobiological concept, specifically for the search for life beyond Earth. Over 1.2 million species have been identified on Earth, yet all life partakes in certain processes, such as homeostasis and replication. Furthermore, several aspects of biochemistry on Earth are thought to be universal, such as the use of organic macromolecules like proteins and nucleic acids. The presence of many biochemical features in empirical data, however, has never been thoroughly investigated. Moreover, the ability to generalize universal features of Earth biology to other worlds suffers from the epistemic problem of induction. Systems biology approaches offer means to quantify abstract patterns in living systems which can more readily be extended beyond Earth’s familiar planetary context. In particular, scaling laws, which characterize how a system responds to changes in size, have met with prior success in investigating universal biology.
This thesis rigorously tests the hypothesis that biochemistry is universal across life on Earth. The study collects enzyme data for annotated archaeal, bacterial, and eukaryotic genomes, in addition to metagenomes. This approach allows one to quantitatively define a biochemical system and sample across known biochemical diversity, while simultaneously exploring enzyme class scaling at both the level of both individual organisms and ecosystems. Using the Kyoto Encyclopedia of Genes and Genomes (KEGG) and the Joint Genome Institute’s Integrated Microbial Genomes and Microbiomes (JGI IMG/M) database, this thesis performs the largest comparative analysis of microbial enzyme content and biochemistry to date. In doing so, this thesis quantitatively explores the distribution of enzyme classes on Earth and adds constraints to notions of universal biochemistry on Earth.
This thesis rigorously tests the hypothesis that biochemistry is universal across life on Earth. The study collects enzyme data for annotated archaeal, bacterial, and eukaryotic genomes, in addition to metagenomes. This approach allows one to quantitatively define a biochemical system and sample across known biochemical diversity, while simultaneously exploring enzyme class scaling at both the level of both individual organisms and ecosystems. Using the Kyoto Encyclopedia of Genes and Genomes (KEGG) and the Joint Genome Institute’s Integrated Microbial Genomes and Microbiomes (JGI IMG/M) database, this thesis performs the largest comparative analysis of microbial enzyme content and biochemistry to date. In doing so, this thesis quantitatively explores the distribution of enzyme classes on Earth and adds constraints to notions of universal biochemistry on Earth.
ContributorsGagler, Dylan (Author) / Walker, Sara I (Thesis advisor) / Kempes, Chris (Committee member) / Trembath-Reichert, Elizabeth (Committee member) / Arizona State University (Publisher)
Created2020