Biocatalytic multi-step production of alkaloids

Alkaloids are natural substances, mostly of complex structure containing several chiral centers. Therefore biocatalytical processes are particularly suited to produce alkaloids due to their high chemo-, regio- and stereoselectivity, and thus outperform traditional chemical catalysis. Generally, significantly higher optical purities are accessible via enzymatic synthesis. Enzymes may be applied in isolated form or as whole cells. Whereas isolated enzymes can be quite costly, the complex metabolism of whole cells may interfere negatively with the respective synthesis reaction. As an alternative, metabolically inactive cellular envelopes containing the respective enzyme anchored in their membranes are worth considering. This can be achieved via a membrane anchor fused to the enzyme to form a fusion protein, which can be inserted post-translationally into the E. coli cytoplasma membrane. Subsequently, a stable pore in the cell wall is created by temperature induced expression of the lysis gene E from the PhiX174 phage. All soluble compounds are expelled from the cell into the medium through this pore, which results in empty cellular envelopes containing the immobilized enzymes.

This project focusses on the establishment of a multi-step biotransformation starting from dopamine to synthesize norlaudanosoline (tetrahydropapaveroline) in bacterial cellular envelopes. To achieve this goal, cellular envelopes with high enzyme activity and enantioselectivity will be developed. Several process variants involving either cellular envelopes in suspension or immobilized cellular envelopes are then compared. The ultimate goal in this project is the establishment of a complete and optimized process capable of biocatalytic synthesis of the chiral norlaudanosoline in its (S)-configuration at the liter scale.