Screening of microorganism producing cephalosporin-C deacetylase and gene cloning = Cephalosporin-C Deacetylase 생산균주의 탐색과 유전자 클로닝

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Cephalosporin-C (CPC) deacetylase [EC 3.1.1.41] (CAH; systemic name, cephalosporin-C acetylhy-drolase) plays a role in deacetylation of C-3 of CPC antibiotics. In this study we obtained the various lipase secreting microorganism by using a tributyrin agar plate method. Visible clear zones due to lipase microorganism, Pseudomonas, having the activity of CPC deacetylase by means of thin layer chromatography (TLC). Supernatants of candidate bacteria culture were reacted with cephalosporin-C and then spotted cellulose plate and was developed with mixture of isopropanol and water (ratio 7 to 3). The CPC deacetylase gene was cloned in E.coli JM109 and sequenced. The cloned DNA(pUCLip) was 2.6kb in length. The nucleotide sequence revealed the presence of one major open reading frame of 1905 nucleotides and encoded a polypeptide consisting of 634 amino acids. The initiation codon of cephalosporin-C deacetylase gene was ATG. The G+C content of the enzyme was 62%. The deduced amino acid sequence showed 58% identity to EstA of Pseuomonas aeruginosa. The consensus sequence maintained among other microorganism cephalosporin-C deacetylases, Gly-X-Ser-X-Gly, was not observed in this protein. Instead cephalosporin-C deacetylase contained the common sequence Gly-Asp-Ser-Leu found in the GDSL family of lipolytic enzymes.
Advisors
Byun, Si-Myung변시명
Description
한국과학기술원 : 생물과학과,
Publisher
한국과학기술원
Issue Date
1999
Identifier
151583/325007 / 000973559
Language
eng
Description

학위논문(석사) - 한국과학기술원 : 생물과학과, 1999.2, [ vi, 51 p. ]

Keywords

Deacetylase; Cephalodporin; Gene cloning; 유전자 클로닝; 디아세틸라아제; 세팔로스포린

URI
http://hdl.handle.net/10203/27882
Link
http://library.kaist.ac.kr/search/detail/view.do?bibCtrlNo=151583&flag=dissertation
Appears in Collection
BS-Theses_Master(석사논문)
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