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Monheim am Rhein, Germany

Richter N.,Evocatal GmbH | Groger H.,Friedrich - Alexander - University, Erlangen - Nuremberg
Applied Microbiology and Biotechnology | Year: 2011

A recombinant enoate reductase from Gluconobacter oxydans was heterologously expressed, purified, characterised and applied in the asymmetric reduction of activated alkenes. In addition to the determination of the kinetic properties, the major focus of this work was to utilise the enzyme in the biotransformation of different interesting compounds such as 3,5,5-trimethyl-2-cyclohexen-1,4-dione (ketoisophorone) and (E/Z)-3,7-dimethyl- 2,6-octadienal (citral). The reaction proceeded with excellent stereoselectivities (>99% ee) as well as absolute chemo- and regioselectivity, only the activated C=C bond of citral was reduced by the enoate reductase, while non-activated C=C bond and carbonyl moiety remained untouched. The described strategy can be used for the production of enantiomerically pure building blocks, which are difficult to prepare by chemical means. In general, the results show that the investigated enoate reductase is a promising catalyst for the use in asymmetric C=C bond reductions. © 2010 Springer-Verlag. Source


Richter N.,Evocatal GmbH | Hummel W.,Heinrich Heine University Dusseldorf
Enzyme and Microbial Technology | Year: 2011

A gene encoding an NADPH-dependent carbonyl reductase from Neurospora crassa (nccr) was cloned and heterologously expressed in Escherichia coli. The enzyme (NcCR) was purified and biochemically characterised. NcCR exhibited a restricted substrate spectrum towards various ketones, and the highest activity (468. U/mg) was observed with dihydroxyacetone. However, NcCR proved to be very selective in the reduction of different α- and β-keto esters. Several compounds were converted to the corresponding hydroxy ester in high enantiomeric excess (ee) at high conversion rates. The enantioselectivity of NcCR for the reduction of ethyl 4-chloro-3-oxobutanoate showed a strong dependence on temperature. This effect was studied in detail, revealing that the ee could be substantially increased by decreasing the temperature from 40 °C (78.8%) to -3 °C (98.0%). When the experimental conditions were optimised to improve the optical purity of the product, (S)-4-chloro-3-hydroxybutanoate (ee 98.0%) was successfully produced on a 300. mg (1.8. mmol) scale using NcCR at -3 °C. © 2011 Elsevier Inc. Source


Grant
Agency: Cordis | Branch: FP7 | Program: CP-FP | Phase: KBBE-2008-3-2-07 | Award Amount: 4.98M | Year: 2009

The Project aims at the mining of individual enzymes and metabolic pathways from extremophilic marine organisms and the metagenomes from microbial communities from peculiar marine environments and consequent funnelling the new enzymatic reactions and processes towards the new biotechnological applications. Project builds up on the scientific and technological excellence of individual academic and industrial partners, and beyond that, on application of the state-of-the-art technologies for archiving, molecular screening for the activities (using a unique Surface Plasmon Resonance screening platform), protein structure elucidation, enzyme engineering and directed evolution and establishing new biotechnological processes (biocatalysis, synthesis of fine chemicals, etc.). Marine sampling hotspots to produce the metagenomic resources for their further exploration will cover the whole diversity of marine microbial life at its limits (hypersaline, low and high temperature, high pressure and low water activity conditions, etc.). Individual enzymes interacting with the substrates will be identified, and in case they are new, hyperexpressed and crystallized and their structures will be elucidated. Consequently, the most promising candidates will be scored against the chiral substrates of relevance for biocatalysis and their ability to perform in water-free systems will be evaluated, the directed evolution will be implemented to improve the performance, and specificity of the enzymes. A comprehensive bioinformatic survey throughout the whole tree of cellular life will reveal and suggest the new candidates homologous to the discovered new proteins, from other organisms to be cloned and assayed. The implementation of the set of new enzymes in the biotechnological processes for fine chemical synthesis and drug discovery will be conducted in a strong alliance with competent industrial partners.


Patent
Evocatal GmbH | Date: 2011-08-16

The present invention relates to the use of a protein comprising an LOV domain for the photosensitive defunctionalization of a molecule and to a method for the photosensitive defunctionalization of a target molecule.


Grant
Agency: Cordis | Branch: FP7 | Program: CP-TP | Phase: KBBE.2011.3.3-02 | Award Amount: 7.77M | Year: 2011

The objective of KYROBIO project is to broaden the toolbox of single enantiomer chiral chemicals that are produced by industry in Europe using biotechnological routes. The main target is applications of lyase enzymes to selectively synthesize molecules with multiple chiral centres applying enzymatic carbon-carbon and carbon-nitrogen bond formation as the key technical platforms. We will then apply synthetic biology to improve fermentation processes in order to generate better enzymes. Chiral compounds are an important class of chemicals that biocatalytic transformation has already demonstrated great potential to compete with chemocatalysts in their production with associated benefits that come from reductions in use of organic solvents, toxic metals and energy but application has been relatively limited. KYROBIO will address the main challenges with moving forward to the next generation of added value industrial applications of white biotechnology for high value chemical synthesis. Using a supradisciplinary approach ranging from enzyme development, chemistry, molecular biology, fermentation and innovative isolation techniques the bottlenecks to applying this new technology will be overcome. It is expected that promising candidate chemicals will be commercialised within three years of completion and so scale up with economic and feasibility studies that are also key technology developments. The consortium includes a strong presence of SMEs including SME leadership and also a large multinational company which ensures multiple routes to market for the outcomes of this project. We will also have economic and life cycle analysis coupled with significant dissemination plans to ensure wider understanding of this technology that will lead to increased acceptance and uptake. The use of this environmentally beneficial technology will help to keep the European chemicals industry at the forefront of white biotechnology and increase opportunities in economic and employment.

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