Asymmetric Stetter reactions catalyzed by thiamine diphosphate-dependent enzymes
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The intermolecular asymmetric Stetter reaction is an almost unexplored transformation for biocatalysts. Previously reported thiamine diphosphate (ThDP)-dependent PigD from Serratia marcescens is the first enzyme identified to catalyze the Stetter reaction of α,β-unsaturated ketones (Michael acceptor substrates) and α-keto acids. PigD is involved in the biosynthesis of the potent cytotoxic agent prodigiosin. Here, we describe the investigation of two new ThDP-dependent enzymes, SeAAS from Saccharopolyspora erythraea and HapD from Hahella chejuensis. Both show a high degree of homology to the amino acid sequence of PigD (39 and 51 %, respectively). The new enzymes were heterologously overproduced in Escherichia coli, and the yield of soluble protein was enhanced by co-expression of the chaperone genes groEL/ES. SeAAS and HapD catalyze intermolecular Stetter reactions in vitro with high enantioselectivity. The enzymes possess a characteristic substrate range with respect to Michael acceptor substrates. This provides support for a new type of ThDP-dependent enzymatic activity, which is abundant in various species and not restricted to prodigiosin biosynthesis in different strains. Moreover, PigD, SeAAS, and HapD are also able to catalyze asymmetric carbon–carbon bond formation reactions of aldehydes and α-keto acids, resulting in 2-hydroxy ketones.
Keywords1,4-Carboligation Biocatalysis C–C coupling Chemoenzymatic synthesis Umpolung
This work was supported by the Deutsche Forschungsgemeinschaft (DFG) in the scope of the Research Group FOR 1296. We are grateful to Prof. Peter Leadlay, University of Cambridge, and Dr. Wolfgang Hüttel, Albert-Ludwigs-Universität Freiburg, for providing plasmid pL1SL2 and for helpful discussions. The technical assistance of Fabrizio Bonina, Volker Brecht, Wolfgang Kornberger, and Simon Waltzer, Albert-Ludwigs-Universität Freiburg, is gratefully acknowledged. We thank Dr. Mostafa Zarei, Center for Biological Systems Analysis, Freiburg, for MS determinations.
- Dresen C (2008) α,β-ungesättigte Carbonyle als Substrate für asymmetrische C-C-Additionen mit Thiamindiphosphat (ThDP)-abhängigen Enzymen. Dissertation, Albert-Ludwigs-Universität FreiburgGoogle Scholar
- Dünnwald T, Demir AS, Siegert P, Pohl M, Müller M (2000) Enantioselective synthesis of (S)-2-hydroxy propanone derivatives by benzoylformate decarboxylase catalyzed C-C-bond formation. Eur J Org Chem 2161–2170Google Scholar
- Enders D, Han J, Henseler A (2008) Asymmetric intermolecular Stetter reactions catalyzed by a novel triazolium derived N-heterocyclic carbene. Chem Commun 3989–3991. doi: 10.1039/B809913H
- Lee JS, Kim Y, Park S, Kim J, Kang S, Lee M, Ryu S, Choi JM, Oh T, Yoon J (2011) Exceptional production of both prodigiosin and cycloprodigiosin as major metabolic constituents by a novel marine bacterium, Zooshikella rubidus S1-1. Appl Environ Microbiol 77:4967–4973. doi: 10.1128/AEM.01986-10 PubMedCentralPubMedCrossRefGoogle Scholar
- Lehwald P (2010) Biokatalytische Synthese tertiärer Alkohole mittels asymmetrischer Carboligationsreaktion unter Verwendung eines Thiamindiphosphat-abängigen Enzyms. Dissertation, Albert-Ludwigs-Universität FreiburgGoogle Scholar
- Williamson NR, Simonsen HT, Ahmed RAA, Goldet G, Slater H, Woodley L, Leeper FJ, Salmond GPC (2005) Biosynthesis of the red antibiotic, prodigiosin, in Serratia: identification of a novel 2-methyl-3-n-amyl-pyrrole (MAP) assembly pathway, definition of the terminal condensing enzyme, and implications for undecylprodigiosin biosynthesis in Streptomyces. Mol Microbiol 56:971–989. doi: 10.1111/j.1365-2958.2005.04602.x PubMedCrossRefGoogle Scholar