A generic biocatalytic reaction platform for the stereospecific reduction and oxidation of alkenes

We propose an ambitious, interdisciplinary research programme to develop a viable manufacturing process, independent of redox coenzymes, to exploit a wide range of novel and industrially relevant redox biotransformations. Our approach is based on the development of (i) microfabricated electrode systems and arrayed microfluidic reactors, (ii) specifically tailored and robust redox catalysts and (iii) the integration and process optimisation of 'engineered catalysts' within the microreactor environment, incorporating on-line quantification of reaction products and facile removal of the desired product by microfluidic liquid-liquid extraction. The technology platform is designed to accommodate a wide range of redox biocatalysts by employing generic methods of electron delivery, and to by-pass the requirement for coenzyme recycling/regeneration. The outcome will be a viable manufacturing process for the production and rapid recovery of high valuable enantiomeric products for the pharmaceutical and synthetic chemistry communities. For the first time, our programme will drive the diverse catalytic potential of redox enzymes towards industrial exploitation, and will thus make a substantial contribution to the field of biocatalysis. The proposed programme is highly innovative and provides generic solutions to the key problems associated with the exploitation of redox enzymes in industrial biocatalytic processes. The programme capitalises on the unique combination of expertise available within the UK Centre of Excellence in Biocatalysis, Bioprocessing and Biomanufacturing (CoEBio3) at the University of Manchester and the excellent research infrastructure available to this group of research workers in the Manchester Interdisciplinary Biocentre. This direct involvement of the 13 member companies of CoEBio3 will provide an invaluable route for effective dissemination and implementation of the project findings.

Redox biotransformations should provide many new, versatile routes to manufacture both chiral and non-chiral speciality chemicals. In practice, it is extremely difficult to operate these reactions economically at scale, since the reactions depend on the provision of stoichiometric quantities of NAD(P)H. Since the cofactors are extremely expensive, economic viability depends on recycling the cofactors. In vitro cofactor recycling using coupled enzymes is generally impractical at large scale (although there are a few exceptions), and alternative approaches, such as artificial cofactors or chemical, electrochemical or photochemical reduction of cofactors, are not industrially viable. As a result, nearly all redox biotransformations are operated using whole cells, exploiting their in-built capacity for cofactor recycling. Even this approach suffers from problems, since cells are notoriously prone to poisoning by substrate/product toxicity, and side reactions may consume substrate or product. Redox biotransformations would be a much more attractive prospect if there were general methods to use isolated enzymes instead of whole cells. In this application, we propose to develop robust, cofactor-independent systems to deliver reducing power to redox enzymes in vitro using microfabricated electrochemical reactors which offer the prospect of a realistic approach to bioelectrocatalytic manufacturing. Our approach is to construct a generic platform technology based on a small number of well characterised catalytic frameworks that enables tailoring of reaction and substrate specificity to target reagents. The platform technology will provide a 'step change' in biocatalysis and allow us to develop a viable manufacturing process, independent of redox coenzymes, to exploit a wide range of novel and industrially relevant redox biotransformations.