Journal
BIOTECHNOLOGY AND BIOENGINEERING
Volume 105, Issue 4, Pages 697-704Publisher
WILEY
DOI: 10.1002/bit.22582
Keywords
7,3 ',4 '-trihydroxyisoflavone (3 ' ODI); daidzein; cytochrome P450; ferredoxin; ferredoxin reductase; recombinant Streptomyces avermitilis MA4680
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Funding
- Korea Science and Engineering Foundation [ROA-2007-000-10007-0]
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Regiospecific 3'-hydroxylation reaction of daidzein was performed with CYP105D7 from Streptomyces avermitilis MA4680 expressed in Escherichia coli. The apparent K-m and k(cat) values of CYP105D7 for daidzein were 21.83 +/- 6.3 mu M and 15.01 +/- 0.6 min(-1) in the presence of 1 mu M of CYP105D7, putidaredoxin (CamB) and putidaredoxin reductase (CamA), respectively. When CYP105D7 was expressed in S. avermitilis MA4680, its cytochrome P450 activity was confirmed by the CO-differente spectra at 450 nm using the whole cell extract. When the whole-cell reaction for the 3'-hydroxylation reaction of daidzein was carried out with 100 mu M of daidzein in 100 mM of phosphate buffer (pH 7.5), the recombinant S. avermitilis grown in R2YE media overexpressing CYP105D7 and ferredoxin FdxH (SAV7470) showed a 3.6-fold higher conversion yield (24%) than the corresponding wild type cell (6.7%). In a 7 L (working volume 3 L) jar fermentor, the recombinants S. avermitilis grown in R2YE media produced 112.5 mg of 7,3',4'-trihydroxyisoflavone (i.e., 29.5%, conversion yield) from 381 mg of daidzein in 15 h. Biotechnol. Bioeng. 2010;105: 697-704. (C) 2009 Wiley Periodicals Inc.
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