원문정보
Cloning and Expression of Inositol Monophosphatase Gene from Streptomyces coelicolor A(3)2
초록
영어
Mycothiol (MSH), a low molecular antioxidant thiol compound, was purified and analyzed from Streptomyces coelicolor A(3)2 by the monobromobimane fluorescence detection method modified by this lab. Through HPLC chromatpgram, MSH fraction was obtained following the elution time of standard MSH (donated by Dr. Robert C. Fahey). That MSH showed the highest
concentration among the thiol compounds contained in the cell indicated that MSH was the key thiol compound having antioxidant activity. To understand the role of gene of inositol monophosphatase (I-1-Pase) involved in the MSH biosynthesis, it was isolated from S. coelicolor A(3)2 and cloned and overexpressed in the Escherichia coli. The expressed I-1-Pase was purified through Ni-NTA column. The soluble protein consisted of 281 amino acids, and the molecular weight was 32 kDa.
I-1-Pase of S. coelicolor A(3)2 had the sequence homology with those of human and E. coli by 24 and 25%, respectively, and had two conserved domains (mofif A and motif B) which were typical of I-1-Pase.
목차
서론
재료 및 방법
사용균주 및 배양
DNA 조작
HPLC chromatogram을 이용한 MSH 분석
Inositol-1-phosphatase 유전자의 클로닝과 발현
발현된 inositol-1-phosphatase 정제
결과 및 고찰
세포내 thiol의 분석
Inositol-1-Phosphatase 유전자의 클로닝 및 발현
I-1-Pase의 아미노산 서열 비교
REFERENCES