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Projects team 'Synthetic enzyme cascades'

Helmholtz young investigators group `Synthetic enzyme cascades´

In focus of our research is the development of value generating processes using synthetic enzyme cascades. We combine catalysts to multi-step biotransformations which do not occur conjunct in nature. Rational enzyme engineering, reaction optimisation and process design is used to gain both, economically and ecologically efficient cascades.
Starting from inexpensive material, highly priced products with direct pharmaceutical application or as building blocks for pharmaceuticals can be yielded. In order to access broad platforms of optically active and highly pure products, we built up toolboxes encompassing enzymes and chemical catalysts with varying substrate preferences and complementary stereo- and regioselectivities. By flexible combinations of these (bio)catalysts can rapidly act on challenging scientific issues with industrial relevance. By doing so, product platforms of hydroxy ketones, amines, amino alcohols and tetrahydroisoquinolines (e.g. latter show a great potential as antibiotics and anticancer drugs) could be established. In some cases, not only in lab scale but also in industrial scale with collaboration partners from industry.

Grafic Micro-aqueous reaction systemMicro-aqueous reaction system

An interdisciplinary team of (bio-) chemists, biologists and engineers develops multi-step biotransformations to chiral products with high step- and atom efficient avoiding (toxic) by-products. Beside multi-step syntheses with purified enzymes and cell extracts in buffered systems, we also investigate the potential of whole cell biotransformations in unconventional media. In focus are micro-aqueous reaction systems or even neat substrate systems. Due to immensely increased substrate concentrations in these non-buffered systems, high product concentrations and space-time-yields can be combined with low production costs. In addition, cheap catalyst and cofactor supply and a facilitated downstream processing renders the application of synthetic enzyme cascades in micro-aqueous reaction systems advantageous.

Current and future research targets are (i) the use of continuous systems and neat substrate systems to further increase space-time-yields, (ii) the effective combination of chemo- and biocatalysts in cascades (iii) setting-up hybrid systems (of chemo-enzymatic cascades and microbial cell factories) to generate value from renewable resources, (iv) regulation of enzyme activities in cascades using light.

Research activities take place at Forschungszentrum Jülich GmbH while teaching activities are conducted at RWTH Aachen University (Aachen Biology and Biotechnology).


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Projects and Cooperations

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Cascade reactions for the synthesis of pharmaceutically relevant substituted amino alcohols and tetrahydroisoquinolines

PhD-student: Vanessa Erdmann

Chiral vicinal amino alcohols are pharmaceutically active compounds or chemical building blocks with various industrial applications. In previous studies all four stereoisomers of the sympathomimetic amino alcohol nor(pseudo)ephedrine could be achieved in a two-step cascade reaction using ThDP-dependent carboligases and amine transferases with high conversions and selectivities. Now this concept is transferred to substituted amino alcohols. Furthermore, formed amino alcohols are the basis for follow-up cyclisation towards tetrahydroisoquinolines using either a third enzyme or a chemical catalyst.

cooperation partners: Wolfgang Kroutil (University of Graz, AT), Helen C. Hailes (University College London, UK), John M. Ward (University College London, UK)

funded by:

Grafic of Helmholtz-Gemeinschaft

Helmholtz Young Investigators Group `Synthetic enzyme cascades´

publication: Erdmann V, Lichman B R, Zhao J, Simon R C, Kroutil W, Ward J M, Hailes H C, Rother D. 2017. Enzymatic and chemoenzymatic 3-step cascades for the synthesis of stereochemically complementary trisubstituted tetrahydroisoquinolines. Angew. Chem. Int. Ed. DOI: 10.1002/anie.201705855

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Catalytic promiscuity of thiamine enzymes – rational combination of structural elements and exploration of the sequence-function space

PhD-student: Saskia Bock

In this DFG Research Group project we want to make use of the full potential of ThDP-based biocatalysis. The consortium collected in-depth knowledge of the enzymes reaction mechanism and the active sites architecture over the last years. Together with recent insights on optimal reaction- and process parameters, we are now able to rationally design optimised catalysts and apply them for industrially feasible carboligation reactions. In a second part of the project the potential of the recently established ThDP-dependent enzyme database is explored by making use of the possibility to walk through the sequence space and investigate and characterise enzymes in detail.

cooperation partners: Jürgen Pleiss (University of Stuttgart), Michael Müller (University of Freiburg)

funded by:

Logo Deutsche Forschungsgemeinschaft

Grafic Thiamine

DFG Research Group FOR 1296

http://www.cofactor-diversity.uni-freiburg.de/FOR1296/FORstart

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Synthetic enzyme cascades towards vicinal amino alcohols and diols from renewables

PhD-student: Kevin Mack

Together with collaboration partners, in this project a complex hybrid example process for the combined production of high-value compounds, starting from D-xylose is investigated. Microbial cell factories will be used converting D-xylose to the starting materials needed for a subsequent biotransformation using synthetic enzyme cascades. This work particularly focuses on investigating the most effective process design for a hydride process combining microbial cell factories and subsequent multi- step enzyme catalysis to gain high valuable vicinal diols and amino alcohols.
In a side project the potential of oxidoreductases for the synthesis of lactones from renewables in neat substrate systems is evaluated.

cooperation partners: Selin Kara (Technical University Hamburg-Harburg), Andrij Pich (RWTH Aachen University), Stephan Noack (FZJ)

funded by:

Grafic of Helmholtz-Gemeinschaft

Grafic BioSC

Helmholtz Young Investigators Group `Synthetic enzyme cascades´

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Reaction and process-optimization of modular synthetic enzyme cascades towards diols and amino alcohols meeting industrial demand

PhD-student: Reinhard Oeggl

Enzymatic cascades have gained increased attention with respect to the development of more sustainable production processes. In this project we aim to address enzymatic processes on different production levels: (i) development of cheap, reproducible, heterologous biocatalyst production via high cell density cultivation, (ii) optimisation of space-time-yields by process engineering– in detail the transition from batch to fed-batch and ultimately continuous production processes– and (iii) integration of in situ product removal.

cooperation partners: Andreas Jupke (RWTH Aachen University), Francesco Molinari and Diego Romano (University of Milan, IT), Stephan Noack (FZJ), Eric von Lieres (FZJ)

funded by:

Grafic of Helmholtz-Gemeinschaft

Helmholtz Young Investigators Group `Synthetic enzyme cascades´

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Guest PhD-student: Federica Dall`Oglio, University of Milan, IT

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terminated projects

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Application of imine reductases in unconventional media

Postdoctoral project of Dr. Zaira Maugeri

funded by: Helmholtz Young Investigators Group `Synthetic enzyme cascades´

Grafic of Helmholtz-Gemeinschaft

Selected publications: Maugeri Z and Rother D. 2017. Reductive Amination of Ketones Catalyzed by Whole Cell Biocatalysts Containing Imine Reductases (IREDs). J. Biotechnol. DOI: 10.1016/j.jbiotec.2017.05.015

Maugeri Z and Rother D. 2016. Application of imine reductases (IREDs) in micro-aqueous reaction systems. Adv. Syn. Catal. 358 (17): 2745-50

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Modularization and optimization of enzymatic reactions applying whole cell biocatalysis in micro-aqueous solvent systems

PhD-thesis by Jochen Wachtmeister

Logo Deutsche Forschungsgemeinschaft


funded by:

in frame of DFG Research Training Group “Bionoco” GK 11 66 at RWTH Aachen
http://www.bionoco.rwth-aachen.de

Selected publications: Wachtmeister J, Jakoblinnert A, Kulig J, Offermann H, Rother D. 2014. Whole Cell Teabag Catalysis for the Modularisation of Synthetic Enzyme Cascades in Micro-Aqueous Systems. ChemCatChem. 6 (4): 1051-1058.

Wachtmeister J, Jakoblinnert A, Rother D. 2016. Enzyme cascades in micro-reaction systems: modular combination of whole cell catalysts gives access to all possible stereosiomers of 1,2-diols with high space-time-yields. Org. Process Res. Dev. 20 (19): 1744–1753

Wachtmeister J, Rother D. 2016. Recent advances in whole cell biocatalysis techniques bridging from investigative to industrial scale. Curr. Opin. Biotechnol. 42: 169–177.

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Tailor-made thiamine diphosphate-dependent enzymes for (S)-selective carboligation

PhD-thesis by Robert Westphal

Logo Deutsche Forschungsgemeinschaft


funded by DFG Research Group FOR 1296
http://www.cofactor-diversity.uni-freiburg.de/FOR1296/FORstart

selected publication: Westphal R, Vogel C, Schmitz C, Pleiss J, Müller M, Pohl M, Rother D. 2014. A Tailor-made Chimeric Thiamine Enzyme for the Direct Asymmetric Synthesis of (S)-Benzoins. Angew. Chem. Int. Ed. 53 (35): 9376-9379.

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Influences of non-conventional media on the selectivity of thiamin-diphosphate (ThDP)-dependent enzymes

Logo Deutsche Forschungsgemeinschaft

PhD-thesis by Tina Gerhards


funded by DFG Research Training Group “Bionoco” GK 11 66
http://www.bionoco.rwth-aachen.de

selected publication: Gerhards T, Mackfeld U, Bocola M, von Lieres E, Wiechert W, Pohl M, Rother D. 2012. Influence of organic solvents on enzymatic asymmetric carboligations. Adv. Synth. Catal., 354: 2805-2820.

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Stereoselective synthesis of vicinal diols with enzymatic cascade reactions

Logo of Marie Curie Actions

PhD-thesis by Justyna Katarzyna Kulig


funded by Marie-Curie Actions (www.biotrains.eu)

selected publications: Kulig J, Frese A, Kroutil W, Pohl M, Rother D. 2013. Biochemical characterization of an alcohol dehydrogenase from Ralstonia sp. Biotech. Bioeng. 110 (7): 1838-1848.

Kulig J, Simon R C, Rose C A, Husain S M, Häckh M, Lüdeke S, Zeitler K, Kroutil W, Pohl M, Rother D. 2012. Stereoselective synthesis of bulky 1,2-diols with alcohol dehydrogenases. Catal. Sci. Technol., 2: 1580-1589.

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Development of a biocatalytic production process for (S)-2-hydroxy ketones

Logo of Marie Curie Actions

PhD-thesis by Álvaro Gómez Baraibar


funded by Marie-Curie Actions (www.biotrains.eu)

selected publication: Baraibar A G, von Lieres E, Wiechert W, Pohl M, Rother D. 2014. Effective (S)-α-hydroxy ketone production: a reaction engineering approach. Top. Catal. 57: 401-411

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2-steps in 1-pot: enzyme cascades for the synthesis of chiral vicinal amino alcohols

Grafic CLIB

PhD-thesis by Torsten Sehl


funded by CLIB Graduate Cluster
http://www.clib-graduatecluster.de/?GC-alumnus-Dr-Torsten-Sehl-wins

Industrial scale production of cathine via a synthetic enzyme cascade approach

Grafic Zentrales Innovationsprogramm Mittelstand

Postdoctoral project of Dr. Torsten Sehl


funded by:

selected publications: Sehl T, Hailes H C, Ward J M, Menyes U, Pohl M, Rother D. 2014. Efficient two-step biocatalytic strategies for the synthesis of all nor(pseudo)ephedrine isomers. Green Chem. 16: 3341-3348.

Sehl T, Hailes H C, Ward J M, Wardenga R, von Lieres E, Offermann H, Westphal R, Pohl M, Rother D. 2013. Two Steps in One Pot: Enzyme Cascade for the Synthesis of Nor(pseudo)ephedrine from Inexpensive Starting Materials. Angew. Chem. Int. Ed. 125 (26): 6904-6908.

Sehl T, Bock S, Marx L, Maugeri Z, Walter L, Westphal R, Vogel C, Menyes U, Erhardt M, Müller M, Pohl M, Rother D. 2017. Asymmetric synthesis of (S)-phenylacetylcarbinol–closing a gap in C–C bond formation. Green Chem. 10.1039/C6GC01803C

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Synthetic enzyme cascades in micro-aqueous reaction media: using whole cell approaches to obtain high product concentrations

Grafic of Helmholtz-Gemeinschaft

Postdoctoral-project of Dr. Andre Jakoblinnert


funded by Helmholtz Young Investigators Group `Synthetic enzyme cascades´

selected publications: Jakoblinnert A, Rother D. 2014. A two-step biocatalytic cascade in micro-aqueous medium: using whole cells to obtain high concentrations of a vicinal diol. Green Chem. 16: 3472-3482.

Erdmann V, Mackfeld U, Rother D, Jakoblinnert A. 2014. Enantioselective, continuous (R)- and (S)-2-butanol synthesis: achieving high space-time-yields with recombinant E. coli cells in a micro-aqueous, solvent-free reaction system. J. Biotechnol. 191: 106-112.


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