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Application of Optogenetics Towards Control of Microbial Chemical Production and in Vivo Protein-Protein Interactions
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| 자료유형 | 학위논문 |
|---|---|
| 서명/저자사항 | Application of Optogenetics Towards Control of Microbial Chemical Production and in Vivo Protein-Protein Interactions. |
| 개인저자 | Zhao, Evan Minghao. |
| 단체저자명 | Princeton University. Chemical and Biological Engineering. |
| 발행사항 | [S.l.]: Princeton University., 2019. |
| 발행사항 | Ann Arbor: ProQuest Dissertations & Theses, 2019. |
| 형태사항 | 247 p. |
| 기본자료 저록 | Dissertations Abstracts International 81-04B. Dissertation Abstract International |
| ISBN | 9781085649353 |
| 학위논문주기 | Thesis (Ph.D.)--Princeton University, 2019. |
| 일반주기 |
Source: Dissertations Abstracts International, Volume: 81-04, Section: B.
Advisor: Avalos, Jose L. |
| 이용제한사항 | This item is not available from ProQuest Dissertations & Theses.This item must not be sold to any third party vendors. |
| 요약 | Optogenetics is control. What are the limits of such control? Can the precision of light-sensitive domains be translated effectively into industrially relevant fields? In this thesis, we engineer optogenetic systems for the applications of microbial chemical production and protein purification. We show that the flexibility of controlling protein interactions with light is a powerful tool for optimizing chemical yields and enabling a new method for protein purification that is solvent independent.We first focused on microbial chemical production. Scientists can engineer the metabolisms of microbes to produce fuel, fragrances, plastics, drugs, etc. However, microbes have not evolved to produce the chemicals we desire. Dynamic regulation of enzyme/protein expression in microbes can overcome this limitation by allowing for healthy growth of microbial catalyst and induction of pathway genes for fermentation. We introduce optogenetics as a regulation mechanism for dynamic control in this thesis.We first characterized an optogenetic transcription system in S. cerevisiae (Chapter 2), developed inverted circuits that activate in the dark, and produced chemicals with these systems. We then improve these circuits by tuning circuit parameters and demonstrate the importance of enhanced kinetics of next generation circuits in chemical production (Chapter 3). Finally, we develop amplification circuits that allow for highly tunable optogenetic activation of transcription with minimal amounts of light to allow for stimulation in high density fermentations (Chapter 4). We then turn our focus to the development of photo-sensitive synthetic membraneless organelles in S. cerevisiae(Chapter 5). We show that using these organelle components as tags enables light- dependent control over metabolic flux on a post-translational level.The final part of this thesis (Chapter 6) tackles a major challenge in industrial protein purification. Industrial protein affinity columns rely on low pH or high salt elution of target proteins due to a lack of non-corrosive control over protein binding. To confront this issue, we generate light-sensitive affinity purification columns, the first in vitro application of light-sensitive proteins.Together, this thesis combines synthetic biology and protein engineering to push optogenetics towards the industrial realm. |
| 일반주제명 | Chemical engineering. Bioengineering. Biochemistry. |
| 언어 | 영어 |
| 바로가기 | 
: 이 자료의 원문은 한국교육학술정보원에서 제공합니다. |
