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Projects team 'Synthetic enzyme cascades'

Team `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).

Graphic ABBT RWTH


http://www.biologie.rwth-aachen.de/cms/Biologie/~kdm/Fachgruppe/?lidx=1

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Projects and Cooperations

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Avoiding cross-reactivities in multi-step biocatalysis by enzyme entrapment in photo-switchable microgels

Postdoc-student: Christiane Claaßen

LightCas aims to develop novel methods and applications in the field of synthetic enzyme cascades by a flexible, optimally orthogonal regulation of each enzymatically catalysed step. High concentrations of desired products in complex enzyme cascades should be gained avoiding cross-reactivity. By means of enzyme entrapment in photo-switchable microgels, reversible enzyme activation and deactivation systems will be investigated. By implementing new hydrogel materials, which are each stimulated at different wavelengths, the selective entrapment and control of enzymes compiled in cascades is targeted.

cooperation partners: Andrij Pich (DWI Aachen)

funded by:

Graphic ERC

Graphic LightCas

European Research Council in frame of ERC starting grant 757320 `LightCas´

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Avoiding cross-reactivities in multi-step biocatalysis by light-induced enzyme deactivation

PhD-student: Tim Gerlach

The targeted deactivation of enzymes in whole cells and enzymes as part of synthetic enzyme cascades by chromophore-assisted laser inactivation will be investigated. Selective deactivation of highly overexpressed enzymes in cells can be seminal not only for whole cell biocatalysis in the clinical and research environment, but also for chip-scale reaction systems and living microbial cell factories.

cooperation partners: Thomas Drepper (IMET, University of Düsseldorf)

funded by:

Graphic ERC

Graphic LightCas

European Research Council in frame of ERC starting grant 757320 `LightCas´

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Synthetic enzyme cascades towards pharmaceutical active amino alcohols starting from renewable carboxylic acids

PhD-student: Douglas Weber

Carboxylic acid reductases are promising biocatalysts for the reduction of substituted aromatic carboxylic acids, which can e.g. be obtained from second generation. The aim of this thesis is to close the missing link in a hybrid process from renewables to active pharmaceutical ingredients. Specifically, the process development will be based on a set of three and four step (chemo)-enzymatic cascade reactions aiming for the production of metaraminol and 2-bromophenyl-tetrahydroisoquinolines from starting materials which is gained from renewables by microbial transformations.

cooperation partners: Stephan Noack (FZJ), Margit Winkler (Graz University of Technology), Nick Turner (Manchester Institute of Biotechnology)

funded by:

Grafic BioSC

Graphic Ministry of Innovation, Science and Research of NRW

FocusFund-Project `HyImPAct´of the Bioeconomy Science Center (BioSC)
Ministry of Innovation, Science and Research of NRW
https://www.biosc.de/Hyimpact_en

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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 reference 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.

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

funded by:

Grafic of Helmholtz-Gemeinschaft

Grafic BioSC

Helmholtz W2/W3-programme `Modular 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. This project aims to address enzymatic processes on different production levels for vicinal diols and amino alcohols: (i) development of cheap, reproducible, heterologous biocatalyst production via high cell density cultivation, (ii) optimisation of space-time-yields by process engineering (transition from batch to fed-batch and ultimately continuous production processes), (iii) cascade applications in micro-aqueous reaction systems– and (iv) integration of facilitated downstream processing concepts.

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

funded by:

Grafic of Helmholtz-Gemeinschaft

Helmholtz W2/W3-programme `Modular enzyme cascades´

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terminated projects

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Synthetic enzyme cascades for the synthesis of amino alcohols and benzylisoquinoline alkaloids

PhD-thesis by Vanessa Erdmann

Grafic of Helmholtz-Gemeinschaft

funded by Helmholtz Young Investigators Group `Synthetic enzyme cascades´

selected 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. 56 (41): 12503-12507

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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

funded by DFG Research Group FOR 1296

Logo Deutsche Forschungsgemeinschaft

Grafic Thiamine

selected publication: 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. 19: 380-384

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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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