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Applied and Environmental Microbiology, November 2007, p. 7246-7251, Vol. 73, No. 22
0099-2240/07/$08.00+0     doi:10.1128/AEM.01411-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Engineering of Artificial Plant Cytochrome P450 Enzymes for Synthesis of Isoflavones by Escherichia coli{triangledown}

Effendi Leonard and Mattheos A. G. Koffas*

Department of Chemical and Biological Engineering, The State University of New York, Buffalo, New York

Received 25 June 2007/ Accepted 15 September 2007

Engineered microbes are becoming increasingly important as recombinant production platforms. However, the nonfunctionality of membrane-bound cytochrome P450 enzymes precludes the use of industrially relevant prokaryotes such as Escherichia coli for high-level in vivo synthesis of many functional plant-derived compounds. We describe the design of a series of artificial isoflavone synthases that allowed the robust production of plant estrogen pharmaceuticals by E. coli. Through this methodology, a plant P450 construct was assembled to mimic the architecture of a self-sufficient bacterial P450 and contained tailor-made membrane recognition signals. The specific in vivo production catalyzed by one identified chimera was up to 20-fold higher than that achieved by the native enzyme expressed in a eukaryotic host and up to 10-fold higher than production by plants. This novel biological device is a strategy for the utilization of laboratory bacteria to robustly manufacture high-value plant P450 products.

* Corresponding author. Mailing address: Chemical and Biological Engineering, 303 Furnas Hall, The State University of New York, Buffalo, NY 14260. Phone: (716) 645-2911, ext. 2221. Fax: (716) 645-3822. E-mail: mkoffas{at}eng.buffalo.edu

{triangledown} Published ahead of print on 28 September 2007.

Applied and Environmental Microbiology, November 2007, p. 7246-7251, Vol. 73, No. 22
0099-2240/07/$08.00+0     doi:10.1128/AEM.01411-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.





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