Campus Units
Plant Biology, Biochemistry, Biophysics and Molecular Biology, Roy J. Carver Department of
Document Type
Article
Publication Version
Accepted Manuscript
Publication Date
9-2015
Journal or Book Title
Applied Microbiology and Biotechnology
Volume
99
Issue
18
First Page
7549
Last Page
7558
DOI
10.1007/s00253-015-6496-2
Abstract
The oxygenation reactions catalyzed by cytochromes P450 (CYPs) play critical roles in plant natural products biosynthesis. At the same time, CYPs are one of most challenging enzymes to functionally characterize due to the difficulty of recombinantly expressing these membrane-associated monooxygenases. In the course of investigating rice diterpenoid biosynthesis we have developed a synthetic biology approach for functional expression of relevant CYPs in Escherichia coli. In certain cases activity was observed for only one of two closely related paralogs although it seems clear that related reactions are required for production of the known diterpenoids. Here we report that optimization of the recombinant expression system enabled characterization of not only these previously recalcitrant CYPs, but also discovery of additional activity relevant to rice diterpenoid biosynthesis. Of particular interest, CYP701A8 was found to catalyze 3β-hydroxylation of syn-pimaradiene, which is presumably relevant to momilactone biosynthesis, while CYP71Z6 & 7 were found to catalyze multiple reactions, with CYP71Z6 catalyzing the production of 2α,3α-dihydroxy-ent-isokaurene via 2α-hydroxy- ent-isokaurene, and CYP71Z7 catalyzing the production of 3α-hydroxy-ent-cassadien-2- one via 2α-hydroxy-ent-cassadiene and ent-cassadien-2-one, which may be relevant to oryzadione and phytocassane biosynthesis, respectively.
Copyright Owner
Springer-Verlag Berlin Heidelberg
Copyright Date
2015
Language
en
File Format
application/pdf
Recommended Citation
Kitaoka, Naoki; Wu, Yisheng; Xu, Meimei; and Peters, Reuben J., "Optimization of recombinant expression enables discovery of novel cytochrome P450 activity in rice diterpenoid biosynthesis" (2015). Biochemistry, Biophysics and Molecular Biology Publications. 91.
https://lib.dr.iastate.edu/bbmb_ag_pubs/91
Included in
Biochemistry Commons, Molecular Biology Commons, Plant Biology Commons, Plant Breeding and Genetics Commons
Comments
This is a manuscript of an article published as Kitaoka, N., Wu, Y., Xu, M. et al. Appl Microbiol Biotechnol (2015) 99: 7549. The final publication is available at Springer via http://dx.doi.org/10.1007/s00253-015-6496-2.