{"id":{"repo_id":"mit","oai_identifier":"oai:dspace.mit.edu:1721.1/145060"},"canonical_url":"https://search.dev.ndltd.org/etd/mit/oai:dspace.mit.edu:1721.1/145060","repository":{"repo_id":"mit","name":"MIT","base_url":"https://dspace.mit.edu/oai/request"},"display":{"title":"Applications of Engineered Proteins in Redox Biology and Biomarker Detection Assay Development","abstract":"Engineered proteins are very versatile tools that have been applied in assay development for various purposes. They have been made into genetically encoded biosensors/probes or affinity agents for biomarker detection. This thesis explored a few topics using assays developed with engineered proteins. The genetically encoded hydrogen peroxide generator, D-amino acid oxidase (DAAO), was used to understand the hours-long intracellular hydrogen peroxide (H₂O₂) generation. This study elucidated that the primary respondent of cytosolic H₂O₂ is peroxiredoxin 1 and the H₂O₂ induced apoptosis initiates before the collapse of Prx/Trx/TR antioxidant network. Then, a genetically encoded FRET sensor was used to design a high-throughput screening assay that identified three small-molecule drugs from over 600 compounds that can mediate toxicity through H₂O₂. This thesis also explored the applications of engineered proteins in diagnostic assay development. I engineered binders against various targets for gram-positive and gram-negative pathogenic bacteria, and two of them that have been tested and showed binding to Salmonella whole cells. The engineered binders were also used to develop a SARS-CoV-2 rapid tests. In this project, sikes lab members developed a paper-based assay to detect the SARS-CoV-2 nucleocapsid protein as a team and successfully validated the assay with patient samples. Subsequently, I improved the thermo-stability of the reporter binder protein used in the assay by switching the fusion partner of the binder to a thermally stable protein. I also identified the bottleneck of an epigentotyping assay development and provided insight for future direction.","abstract_html":"Engineered proteins are very versatile tools that have been applied in assay development for various purposes. They have been made into genetically encoded biosensors/probes or affinity agents for biomarker detection. This thesis explored a few topics using assays developed with engineered proteins. The genetically encoded hydrogen peroxide generator, D-amino acid oxidase (DAAO), was used to understand the hours-long intracellular hydrogen peroxide (H₂O₂) generation. This study elucidated that the primary respondent of cytosolic H₂O₂ is peroxiredoxin 1 and the H₂O₂ induced apoptosis initiates before the collapse of Prx/Trx/TR antioxidant network. Then, a genetically encoded FRET sensor was used to design a high-throughput screening assay that identified three small-molecule drugs from over 600 compounds that can mediate toxicity through H₂O₂. This thesis also explored the applications of engineered proteins in diagnostic assay development. I engineered binders against various targets for gram-positive and gram-negative pathogenic bacteria, and two of them that have been tested and showed binding to Salmonella whole cells. The engineered binders were also used to develop a SARS-CoV-2 rapid tests. In this project, sikes lab members developed a paper-based assay to detect the SARS-CoV-2 nucleocapsid protein as a team and successfully validated the assay with patient samples. Subsequently, I improved the thermo-stability of the reporter binder protein used in the assay by switching the fusion partner of the binder to a thermally stable protein. I also identified the bottleneck of an epigentotyping assay development and provided insight for future direction.","abstract_has_math":false,"creators":["Hao, Yining"],"institution":"Massachusetts Institute of Technology","degree_name":"Doctoral","degree_level":null,"degree_discipline":null,"degree_department":"Massachusetts Institute of Technology. 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They have been made into genetically encoded biosensors/probes or affinity agents for biomarker detection. This thesis explored a few topics using assays developed with engineered proteins. The genetically encoded hydrogen peroxide generator, D-amino acid oxidase (DAAO), was used to understand the hours-long intracellular hydrogen peroxide (H₂O₂) generation. This study elucidated that the primary respondent of cytosolic H₂O₂ is peroxiredoxin 1 and the H₂O₂ induced apoptosis initiates before the collapse of Prx/Trx/TR antioxidant network. Then, a genetically encoded FRET sensor was used to design a high-throughput screening assay that identified three small-molecule drugs from over 600 compounds that can mediate toxicity through H₂O₂. This thesis also explored the applications of engineered proteins in diagnostic assay development. I engineered binders against various targets for gram-positive and gram-negative pathogenic bacteria, and two of them that have been tested and showed binding to Salmonella whole cells. The engineered binders were also used to develop a SARS-CoV-2 rapid tests. In this project, sikes lab members developed a paper-based assay to detect the SARS-CoV-2 nucleocapsid protein as a team and successfully validated the assay with patient samples. Subsequently, I improved the thermo-stability of the reporter binder protein used in the assay by switching the fusion partner of the binder to a thermally stable protein. I also identified the bottleneck of an epigentotyping assay development and provided insight for future direction."]},{"key":"dc:description.degree","label":"Dc Description Degree","values":["Ph.D."]},{"key":"dc:title","label":"Title","values":["Applications of Engineered Proteins in Redox Biology and Biomarker Detection Assay Development"]}]}],"canonical_facts":{"dc:contributor.advisor":["Sikes, Hadley D."],"dc:contributor.department":["Massachusetts Institute of Technology. Department of Chemical Engineering"],"dc:creator":["Hao, Yining"],"dc:date.accessioned":["2022-08-29T16:30:15Z"],"dc:date.available":["2022-08-29T16:30:15Z"],"dc:date.issued":["2022-05"],"dc:description.abstract":["Engineered proteins are very versatile tools that have been applied in assay development for various purposes. They have been made into genetically encoded biosensors/probes or affinity agents for biomarker detection. This thesis explored a few topics using assays developed with engineered proteins. The genetically encoded hydrogen peroxide generator, D-amino acid oxidase (DAAO), was used to understand the hours-long intracellular hydrogen peroxide (H₂O₂) generation. This study elucidated that the primary respondent of cytosolic H₂O₂ is peroxiredoxin 1 and the H₂O₂ induced apoptosis initiates before the collapse of Prx/Trx/TR antioxidant network. Then, a genetically encoded FRET sensor was used to design a high-throughput screening assay that identified three small-molecule drugs from over 600 compounds that can mediate toxicity through H₂O₂. This thesis also explored the applications of engineered proteins in diagnostic assay development. I engineered binders against various targets for gram-positive and gram-negative pathogenic bacteria, and two of them that have been tested and showed binding to Salmonella whole cells. The engineered binders were also used to develop a SARS-CoV-2 rapid tests. In this project, sikes lab members developed a paper-based assay to detect the SARS-CoV-2 nucleocapsid protein as a team and successfully validated the assay with patient samples. Subsequently, I improved the thermo-stability of the reporter binder protein used in the assay by switching the fusion partner of the binder to a thermally stable protein. I also identified the bottleneck of an epigentotyping assay development and provided insight for future direction."],"dc:description.degree":["Ph.D."],"dc:identifier.uri":["https://hdl.handle.net/1721.1/145060"],"dc:publisher":["Massachusetts Institute of Technology"],"dc:rights":["In Copyright - Educational Use Permitted","Copyright MIT"],"dc:rights.uri":["http://rightsstatements.org/page/InC-EDU/1.0/"],"dc:title":["Applications of Engineered Proteins in Redox Biology and Biomarker Detection Assay Development"],"dc:type":["Thesis"],"thesis:degree_name":["Doctoral","Doctor of Philosophy"]},"updated_at":"2026-07-22T22:22:02Z"}