Regioselective hydroxylation of daidzein using P450 (CYP105D7) from Streptomyces avermitilis MA4680.

Abstract

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 microM and 15.01 +/- 0.6 min(-1) in the presence of 1 microM 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-difference 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 microM 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.

DOI: 10.1002/bit.22582

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@article{Pandey2010RegioselectiveHO, title={Regioselective hydroxylation of daidzein using P450 (CYP105D7) from Streptomyces avermitilis MA4680.}, author={Bishnu Prasad Pandey and Changhyun Roh and Kwon-Young Choi and Nahum Lee and Eun Jung Kim and Sungghi Ko and Taejin Kim and Hyundon Yun and Byung-Gee Kim}, journal={Biotechnology and bioengineering}, year={2010}, volume={105 4}, pages={697-704} }