DC Field | Value | Language |
---|---|---|
dc.contributor.advisor | Jeong, Ki Jun | - |
dc.contributor.advisor | 정기준 | - |
dc.contributor.author | Bang, Hyun Bae | - |
dc.date.accessioned | 2021-05-12T19:45:46Z | - |
dc.date.available | 2021-05-12T19:45:46Z | - |
dc.date.issued | 2020 | - |
dc.identifier.uri | http://library.kaist.ac.kr/search/detail/view.do?bibCtrlNo=924550&flag=dissertation | en_US |
dc.identifier.uri | http://hdl.handle.net/10203/284461 | - |
dc.description | 학위논문(박사) - 한국과학기술원 : 생명화학공학과, 2020.8,[ix, 132 p. :] | - |
dc.description.abstract | Cinnamaldehyde derived from cinnamon bark has received big attention as a potent nematicide due to its high nematicidal activity. Previously in Master’s thesis, I succeeded in developing Escherichia coli (YHP05 strain) able to synthesize cinnamaldehyde. However, production titer (75 mg/L) was not enough high for commercialization, and it is desired to further engineer host for enhanced production of cinnamaldehyde. First, I engineered two amino acids-auxotrophic YHP05 strain to prototrophic strain for the economic process of cinnamaldehyde production. Next, for the efficient conversion of trans-cinnamic acid to cinnamaldehyde, I constructed a single-step conversion system by introducing co-expression system of carboxylic acid reductase (CAR) and phosphopantetheinyl transferase (PPTase) genes. In addition, to prevent the spontaneous conversion of cinnamaldehyde to cinnamyl alcohol, total 10 endogenous reductases and dehydrogenases genes were deleted. Fourth, after comparison of promoters and integration positions, cinnamaldehyde production system was integrated into optimal positions of E. coli chromosomal DNA for development of antibiotic-free, inducer-free system. Fifth, reinforcement of cofactor generation was performed for enhanced production of cinnamaldehyde. Next, to reduce the inhibitory effect of cinnamaldehyde and improve the production titer, in situ product removal (ISPR) strategy was applied in cultivations. However, cell growth retardation was occurred by acetate accumulation. To solve this problem, acetate production pathway was deleted by newly constructed CRISPR/Cas9-based scarless genome editing system. Additionally, to decrease cell toxicity to cinnamaldehyde, cell recycle system with a polymer resin and promoter replacement of efflux pumps (TolC and AcrAB) were tested. Through these metabolic engineering and process optimization strategies, I successfully demonstrated the gram-scale production of cinnamaldehyde. I also confirmed that cinnamaldehyde produced by our engineered E. coli had a nematicidal activity similar to the activity of authentic cinnamaldehyde by nematicidal assays against Bursaphelenchus xylophilus. | - |
dc.language | eng | - |
dc.publisher | 한국과학기술원 | - |
dc.subject | Escherichia coli▼aCinnamaldehyde▼aNematode▼aMetabolic engineering▼aChromosomal integration▼aCofactor balance▼aEfflux pump▼aFed-batch cultivation▼aTwo-phase partitioning bioreactor (TPPB) | - |
dc.subject | 대장균▼a신남알데하이드▼a선충▼a대사 공학▼a유전자 삽입▼a보조인자 균형▼a유출 펌프▼a세포 고농도 배양▼a이상 분리 배양 | - |
dc.title | Metabolic engineering of Escherichia coli and process optimization for high-level production of cinnamaldehyde | - |
dc.title.alternative | 신남알데하이드의 대량 생산을 위한 대사 공학 기반 대장균주의 개량 및 공정 최적화 | - |
dc.type | Thesis(Ph.D) | - |
dc.identifier.CNRN | 325007 | - |
dc.description.department | 한국과학기술원 :생명화학공학과, | - |
dc.contributor.alternativeauthor | 방현배 | - |
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