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KMID : 0380020040190060462
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Korean Journal of Biotechnology and Bioengineering
2004 Volume.19 No. 6 p.462 ~ p.466
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Cloning and Expression of Inositol Monophosphatase Gene from Streptomyces coelicolor A[3]2
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Kim Jin-Kwon
Choi Hak-Sun
Kim Sung-Joon
Kim Si-Wook
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Abstract
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Mycothiol (MSH), a low molecular antioxidant thiol compound, was purified and analyzed from Streptomyns 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 , respectively, and had two conserved domains (mofif A and motif B) which were typical of I-1-Pase.
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KEYWORD
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Mycothiol, Streptomyces coelicolor A(3)2, monobromobimane fluorescence detection method, inositol monophosphatase (I-1-Pase)
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