Natural products biosynthesis often requires the action of multiple CYPs (cytochromes P450), whose ability to introduce oxygen, increasing solubility, is critical for imparting biological activity. In previous investigations of rice diterpenoid biosynthesis, we characterized CYPs that catalyse alternative hydroxylation of ent-sandaracopimaradiene, the precursor to the rice oryzalexin antibiotic phytoalexins. In particular, CYP76M5, CYP76M6 and CYP76M8 were all shown to carry out C-7β hydroxylation, whereas CYP701A8 catalyses C-3α hydroxylation, with oxy groups found at both positions in oryzalexins A–D, suggesting that these may act consecutively in oryzalexin biosynthesis. In the present paper, we report that, although CYP701A8 only poorly reacts with 7β-hydroxy-ent-sandaracopimaradiene, CYP76M6 and CYP76M8 readily react with 3α-hydroxy-ent-sandaracopimaradiene. Notably, their activity yields distinct products, resulting from hydroxylation at C-9β by CYP76M6 or C-7β by CYP76M8, on different sides of the core tricyclic ring structure. Thus CYP76M6 and CYP76M8 have distinct non-redundant roles in orzyalexin biosynthesis. Moreover, the resulting 3α,7β- and 3α,9β-diols correspond to oryzalexins D and E respectively. Accordingly, the results of the present study complete the functional identification of the biosynthetic pathway underlying the production of these bioactive phytoalexins. In addition, the altered regiochemistry catalysed by CYP76M6 following C-3α hydroxylation has some implications for its active-site configuration, offering further molecular insight.
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Research Article|
August 09 2013
Picking sides: distinct roles for CYP76M6 and CYP76M8 in rice oryzalexin biosynthesis Available to Purchase
Yisheng Wu;
Yisheng Wu
1
*Department of Biochemistry, Biophysics, and Molecular Biology, Iowa State University, Ames, IA 50011, U.S.A.
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Qiang Wang;
Qiang Wang
2
*Department of Biochemistry, Biophysics, and Molecular Biology, Iowa State University, Ames, IA 50011, U.S.A.
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Matthew L. Hillwig;
Matthew L. Hillwig
3
*Department of Biochemistry, Biophysics, and Molecular Biology, Iowa State University, Ames, IA 50011, U.S.A.
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Reuben J. Peters
Reuben J. Peters
4
*Department of Biochemistry, Biophysics, and Molecular Biology, Iowa State University, Ames, IA 50011, U.S.A.
4To whom correspondence should be addressed (email [email protected]).
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Publisher: Portland Press Ltd
Received:
April 24 2013
Revision Received:
June 24 2013
Accepted:
June 25 2013
Accepted Manuscript online:
June 25 2013
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© The Authors Journal compilation © 2013 Biochemical Society
2013
Biochem J (2013) 454 (2): 209–216.
Article history
Received:
April 24 2013
Revision Received:
June 24 2013
Accepted:
June 25 2013
Accepted Manuscript online:
June 25 2013
Citation
Yisheng Wu, Qiang Wang, Matthew L. Hillwig, Reuben J. Peters; Picking sides: distinct roles for CYP76M6 and CYP76M8 in rice oryzalexin biosynthesis. Biochem J 1 September 2013; 454 (2): 209–216. doi: https://doi.org/10.1042/BJ20130574
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