ABSTRACT The ultimate step in the formation of thebaine, a pentacyclic opiate alkaloid readily converted to the narcotic analgesics codeine and morphine in the opium poppy, has long been

presumed to be a spontaneous reaction. We have detected and purified a novel enzyme from opium poppy latex that is capable of the efficient formation of thebaine from (7_S_)-salutaridinol

7-_O_-acetate at the expense of labile hydroxylated byproducts, which are preferentially produced by spontaneous allylic elimination. Remarkably, thebaine synthase (THS), a member of the

pathogenesis-related 10 protein (PR10) superfamily, is encoded within a novel gene cluster in the opium poppy genome that also includes genes encoding the four biosynthetic enzymes

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OTHERS ALKALOID BINDING TO OPIUM POPPY MAJOR LATEX PROTEINS TRIGGERS STRUCTURAL MODIFICATION AND FUNCTIONAL AGGREGATION Article Open access 09 November 2022 ENGINEERING THE AMOEBA

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references ACKNOWLEDGEMENTS We are grateful to L. Brechenmacher and the Southern Alberta Mass Spectrometry Centre for assistance with the proteomics analysis, S. Yeaman for guidance with the

genome assembly, and T. Back for advice on chemical reaction mechanisms. We acknowledge the expert services provided by the McGill University-Genome Québec Innovation Centre with respect to

genome sequencing and preliminary assembly. We also thank A. Pigula, S. Muley, E. Eberhard, A. Kumar, K. Hetenyi, I. Esaid, H. Tang, H. Roth, M. Schmalisch, L. Hom, C. Savile, P.

Seufer-Wasserthal, T. Noh, B. Walsh, and R. J. Kirk for technical assistance and project guidance. This work was supported by Genopaver, LLC., Epimeron Inc. and funds awarded through the

Industrial Research Assistance Program (IRAP; Project 86155) operated by the National Research Council of Canada to Epimeron, Inc. AUTHOR INFORMATION AUTHORS AND AFFILIATIONS * Department of

Biological Sciences, University of Calgary, Calgary, Canada Xue Chen, Jillian M. Hagel, Limei Chang & Peter J. Facchini * Epimeron Inc., Calgary, Canada Xue Chen, Jillian M. Hagel, 

Limei Chang, Joseph E. Tucker & Peter J. Facchini * Department of Biochemistry and Molecular Biology, University of Calgary, Calgary, Canada Joseph E. Tucker * Intrexon Corp., Industrial

Products Division, South San Francisco, CA, USA Stacey A. Shiigi, Yuora Yelpaala, Hsiang-Yun Chen, Rodrigo Estrada, Jeffrey Colbeck, Maria Enquist-Newman, Ana B. Ibáñez, Guillaume Cottarel 

& Genevieve M. Vidanes Authors * Xue Chen View author publications You can also search for this author inPubMed Google Scholar * Jillian M. Hagel View author publications You can also

search for this author inPubMed Google Scholar * Limei Chang View author publications You can also search for this author inPubMed Google Scholar * Joseph E. Tucker View author publications

You can also search for this author inPubMed Google Scholar * Stacey A. Shiigi View author publications You can also search for this author inPubMed Google Scholar * Yuora Yelpaala View

author publications You can also search for this author inPubMed Google Scholar * Hsiang-Yun Chen View author publications You can also search for this author inPubMed Google Scholar *

Rodrigo Estrada View author publications You can also search for this author inPubMed Google Scholar * Jeffrey Colbeck View author publications You can also search for this author inPubMed 

Google Scholar * Maria Enquist-Newman View author publications You can also search for this author inPubMed Google Scholar * Ana B. Ibáñez View author publications You can also search for

this author inPubMed Google Scholar * Guillaume Cottarel View author publications You can also search for this author inPubMed Google Scholar * Genevieve M. Vidanes View author publications

You can also search for this author inPubMed Google Scholar * Peter J. Facchini View author publications You can also search for this author inPubMed Google Scholar CONTRIBUTIONS X.C.,

J.M.H., J.E.T., G.C., G.M.V., and P.J.F. contributed to the project design; X.C. performed most of the enzymology, protein purification, molecular biology, gene silencing, and transcript

profiling work; J.M.H. conducted the high-resolution LC-MS analysis and proposed the reaction mechanism; L.C., S.A.S., and M.E.-N. developed testing conditions and characterized gene

candidates in yeast; R.E. and J.C. designed and built plasmids and yeast strains; Y.Y., H.-Y.C., and A.B.I. developed and performed LC-MS analysis for yeast fermentation experiments; X.C.,

J.M.H., and P.J.F. wrote the manuscript. P.J.F. supervised the project. All authors have read and approved the content of the manuscript. CORRESPONDING AUTHOR Correspondence to Peter J.

Facchini. ETHICS DECLARATIONS COMPETING INTERESTS Patent applications related to this work have been filed (PCT/CA2017/050779 and PCT/US2017/039589). ADDITIONAL INFORMATION PUBLISHER’S NOTE:

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Figures 1–22, Supplementary Tables 1–8, Supplementary Note 1–3 REPORTING SUMMARY RIGHTS AND PERMISSIONS Reprints and permissions ABOUT THIS ARTICLE CITE THIS ARTICLE Chen, X., Hagel, J.M.,

Chang, L. _et al._ A pathogenesis-related 10 protein catalyzes the final step in thebaine biosynthesis. _Nat Chem Biol_ 14, 738–743 (2018). https://doi.org/10.1038/s41589-018-0059-7 Download

citation * Received: 17 October 2017 * Accepted: 16 March 2018 * Published: 28 May 2018 * Issue Date: July 2018 * DOI: https://doi.org/10.1038/s41589-018-0059-7 SHARE THIS ARTICLE Anyone

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