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The molecular architecture of major enzymes from ajmaline biosynthetic pathway

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Abstract

The biosynthetic pathway leading to the monoterpenoid indole alkaloid ajmaline in Rauvolfia serpentiin serpentina is one of the most studied in the field of natural product biosynthesis. Ajmaline has a complex structure which is based on a six-membered ring system harbouring nine chiral carbon atoms. There are about fifteen enzymes involved, including some involving the side reactions of the ajmaline biosynthetic pathway. All enzymes exhibit pronounced substrate specificity. In the recent years isolation and sequencing of their cDNAs has allowed a detailed sequence analysis and comparison with functionally related and occasionally un-related enzymes. Site-directed mutations of several of the ajmaline-synthesizing enzymes have been performed and their catalytic residues have been identified. Success with over-expression of the enzymes was an important step for their crystallization and structural analysis by X-ray crystallography. Crystals with sufficient resolution were obtained from the major enzymes of the pathway. Strictosidine synthase has a 3D-structure with a six-bladed β-propeller fold the first time such a fold found in the plant kingdom. Its ligand complexes with tryptamine and secologanin, as well as structure-based sequence alignment, indicate a possible evolutionary relationship to several primary sequence-unrelated structures with this fold. The structure of strictosidine glucosidase was determined and its structure has as a (β/α)8 barrel fold. Vinorine synthase provides the first 3D structure of a member of BAHD enzyme super-family. Raucaffricine glucosidase involved in a side-route of ajmaline biosynthesis has been crystallized. The ajmaline biosynthetic pathway is an outstanding example where many enzymes 3D-structure have been known and where there is a real potential for protein engineering to yield new alkaloid.

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Acknowledgements

We are very thankful to Mrs. Doris Rohr of our research group for cultivation of all plant cell systems, to Prof. F. Lottspeich and his coworkers Mrs. I. Mathes and Mr. R. Mentele (Max-Planck-Institute of Biochemistry, Martinsried, Germany) for their enthusiastic help in sequencing pre-purified Rauvolfia enzymes and to Prof. H. Michel, Dr. G. Fritzsch and Dr. J. Koepke (Max-Planck-Institute of Biophysics, Frankfurt/Main, Germany) for introducing us to structural biology research.

Our research was also supported by the Deutsche Forschungsgemeinschaft (Bonn, Bad-Godesberg, Germany), Fonds der Chemischen Industrie (Frankfurt/Main, Germany) and the Bundesministerium für Bildung und Forschung (BMBF), Bonn, Germany). Support in the form of access to synchrotron facilities by the European Community (Research Infrastructure Action under the FP6 “Structuring the European Research Area Programme”, contact number RII3/CT/2004/5060008) is also␣acknowledged and we thank Dr. Paul Tucker (EMBL Hamburg, Germany) for critically reading the manuscript.

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Authors and Affiliations

  1. Department of Pharmaceutical Biology, Institute of Pharmacy, Johannes Gutenberg-Universität, Staudinger Weg 5, D-55099, Mainz, Germany

    Joachim Stöckigt, Martin Ruppert, Leif Barleben, Xueyan Ma, Elke Loris & Marco Hill

  2. European Molecular Biology Laboratory Hamburg Outstation Deutsches Elektronen-Synchrotron, Notkestrasse 85, D-22603, Hamburg, Germany

    Santosh Panjikar

  3. College of Pharmaceutical Sciences, Zhejiang University, 353 Yan An Road, Hangzhou, 310031, China

    Joachim Stöckigt

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  1. Joachim Stöckigt
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  2. Santosh Panjikar
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Correspondence to Joachim Stöckigt.

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Stöckigt, J., Panjikar, S., Ruppert, M. et al. The molecular architecture of major enzymes from ajmaline biosynthetic pathway. Phytochem Rev 6, 15–34 (2007). https://doi.org/10.1007/s11101-006-9016-2

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  • DOI: https://doi.org/10.1007/s11101-006-9016-2

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