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Applied and Environmental Microbiology, November 2008, p. 6908-6917, Vol. 74, No. 22
0099-2240/08/$08.00+0     doi:10.1128/AEM.01495-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Biosynthesis of Ubiquinone Compounds with Conjugated Prenyl Side Chains{triangledown} ,{dagger}

Pyung Cheon Lee,1,3 Christine Salomon,2 Benjamin Mijts,1,{ddagger} and Claudia Schmidt-Dannert1*

Department of Biochemistry, Molecular Biology, and Biophysics,1 Center for Drug Design, University of Minnesota, St. Paul, Minnesota,2 Department of Molecular Science and Technology, Ajou University, Suwon, South Korea3

Received 2 July 2008/ Accepted 15 September 2008

Enzymatic steps from two different biosynthetic pathways were combined in Escherichia coli, directing the synthesis of a new class of biomolecules—ubiquinones with prenyl side chains containing conjugated double bonds. This was achieved by the activity of a C30 carotenoid desaturase, CrtN, from Staphylococcus aureus, which exhibited an inherent flexibility in substrate recognition compared to other carotenoid desaturases. By utilizing the known plasticity of E. coli's native ubiquinone biosynthesis pathway and the unusual activity of CrtN, modified ubiquinone structures with prenyl side chains containing conjugated double bonds were generated. The side chains of the new structures were confirmed to have different degrees of desaturation by mass spectrometry and nuclear magnetic resonance analysis. In vivo 14C labeling and in vitro activity studies showed that CrtN desaturates octaprenyl diphosphates but not the ubiquinone compounds directly. Antioxidant properties of conjugated side chain ubiquinones were analyzed in an in vitro β-carotene-linoleate model system and were found to be higher than the corresponding unmodified ubiquinones. These results demonstrate that by combining pathway steps from different branches of biosynthetic networks, classes of compounds not observed in nature can be synthesized and structural motifs that are functionally important can be combined or enhanced.

* Corresponding author. Mailing address: Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, 1479 Gortner Avenue, St. Paul, MN 55108. Phone: (612) 625-5782. Fax: (612) 625-5780. E-mail: schmi232{at}umn.edu

{triangledown} Published ahead of print on 26 September 2008.

{dagger} Supplemental material for this article may be found at http://aem.asm.org/.

{ddagger} Present address: Codexis, Inc., Redwood City, CA 94063.

Applied and Environmental Microbiology, November 2008, p. 6908-6917, Vol. 74, No. 22
0099-2240/08/$08.00+0     doi:10.1128/AEM.01495-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.





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