Workpackage 3: Transaminase-catalyzed multi-enzyme cascade reactions

Workpackage Leader: Jun.-Prof. Dr. Matthias Höhne
Lead beneficiary: Ernst-Moritz-Arndt-Universität Greifswald
Participating Partners: Entrechem, SARomics, Enzymicals, KTH, Viazym
Start month: 7 End month: 42

Description of work and role of partners

Novel equilibrium shift strategies: ESR 6 investigates novel achiral amine-donors, such as α,ω-diamines, (NH2-(CH2)n-NH2; n=2-5) and ω-aminoalkan-1-ols (NH2-(CH2)n-NH2; n=4-5) and the oxidation of the formed hemiacetal byproduct by a dehydrogenase to efficiently aminate model ketones. Once the best amino-donor was identified in terms of cost and reaction efficiency, different prochiral ketones, hydroxyketones and diketones from cascades 4-7 and of commercial interest will be used in collaboration with Entrechem and Enzymicals.

Synthesis of 1-amino-cyclohexane-4-ol: After a setup of the analytics using commercially available substrates/products,ESR 7 performs a screening of the in-house collection of KREDs and ω-TA to identify enantio and regio-selective enzymes. Special emphasis is given to identify enzymes that minimize possible by-products formation (diol- or diamine).
ESR 7 then explores cascade reactions and integrates the aforementioned strategy for equilibrium shift, As during the oxidation of the hemiacetal co-product NADH is formed, he investigates whether these reduction equivalents are sufficient for the ketone reduction step. Reversible imine formation which might lead to polymerization of the intermediate aminoketone will be addressed by a proper fine-tuning of the reaction conditions (e.g. pH-value, high keto reductase activity to scavenge the amino ketone. Finally, ESR7 will optimize STY and productivity by reaction engineering, and protein engineering, if necessary. Enzyme production will be optimized to facilitate large scale production.

For the development of a dynamic kinetic resolution, ESR9 optimizes a couple of enantiocomplementary transaminases to emulate an amine racemase. Thereby the focus lies on protein engineering and immobilization strategies to increase stability in a biophasic or pure organic solvent system. Multivariate methods will then be used at Saromics to predictbeneficial combinations of mutations. In parallel, a DKR using Penicillin G acylase able to acylate in aqueous media will be investigated.


    1.) Improving the transaminase technology: Expanding the substrate scope and design of novel equilibrium shift strategies.
    2.) Synthesis of optically pure 1-amino-cyclohexane-4-ol by combining enzymatic keto reduction and efficient transamination.
    3.) Synthesis of enantiomerically pure chiral amine derivatives in a deracemization cascade in organic solvent or biphasic reaction media, creation of new stable transaminases based on multivariate methodology and screening data.
    4.) Synthesis of bulky amines by combining Pd-catalyzed Suzuki-crosscoupling and transamination.

The Biocascades Project is a joint collaboration among the following university and industrial partners