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Expression, purification and characterization of ubiquitin-specific protease 1 (UBP1) for hydrolysis of ubiquitin-fused hGH expressed in recombinant E. coli : 재조합 대장균에서 생성된 유비퀴틴 융합 hGH를 가수분해하는 ubiquitin specific protease 1 (UBP1)의 발현, 정제 및 효소특성에 관한 연구
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- Authors
- Advisor
- 서진호
- Issue Date
- 2003
- Publisher
- 서울대학교 대학원
- Keywords
- ubiquitin-specific protease 1 (UBP1) ; Recombinant E. coli ; 재조합 대장균 ; P. pastoris ; 피키아 파스토리스 ; Polycationic fusion tails ; 양이온 융합 꼬리 ; Fusion partners ; 융합 파트너 ; High cell density cultivation technology ; 고농도 배양 ; Immobilized metal affinity chromatography (imac) ; 고정화 금속 친화 크로마토그래피 ; Ubiquitin-hgh (human growth hormone)
- Description
- Thesis (master`s)--서울대학교 대학원 :농생명공학부,2003.
- Abstract
- This research was focused on expression, purification and characterization of ubiquitinspecific
protease 1 (UBP1) expressed in recombinant E. coli and P. pastoris. Various
systems were constructed by fusing polycationic fusion tails or fusion partners to the C- or
N-terminus of the product protein. In particular, UBP1 containing 6 histidine residues at
the N-terminal end showed best results in terms of expression level and purification
efficiency.
UBP1 expressed in P. pastoris was secreted to the medium and enzyme activity was
detected after 48 hours induction with methanol. The N-terminal 6×His-tagged UBP1 was
overproduced in recombinant E. coli using high cell density cultivation technology and
purified using immobilized metal affinity chromatography (IMAC). The molecular weight
of UBP1 was found to be 83,500 daltons. The optimum temperature and pH for the
enzyme reaction when ubiquitin-hGH (human growth hormone) was used as a substrate
were 40℃ and pH 8.0, respectively.
- Language
- English
- URI
- http://dcollection.snu.ac.kr:80/jsp/common/DcLoOrgPer.jsp?sItemId=000000057504
https://hdl.handle.net/10371/68522
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