Campus Units

Biochemistry, Biophysics and Molecular Biology, Plant Biology

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.

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.

Copyright Owner

Springer-Verlag Berlin Heidelberg

Language

en

File Format

application/pdf

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