FWF - Active-site design - Active-site design of cofactor-free enzymes

Project: Research project

Project Details


Enzymes are the catalysts of Nature. Their mild reactions conditions and high selectivity make them ideal tools for organic synthesis. Many biocatalysts, however, are characterized by limited substrate spectra. This requires methods for the modification of enzymatic mechanisms in order to create new reactivities. The breaking and formation of bonds between carbon atoms is of high interest in view of synthetic applications. However, these reactions are very challenging for biocatalysis. The bacterial enzymes arylmalonate decarboxylase and eudesmol synthase catalyse these reactions with outstanding selectivity. The molecular mechanisms of this selectivity lie in the selective stabilization of reaction intermediates on the one hand, and in targeted quenching of these intermediates on the other hands. On basis of mechanistic studies and computational simulations, we aim to incorporate functional groups at specific positions in order to direct the reactions of both enzymes towards the formation of new products. With the current knowledge, it is extremely difficult to formulate precise predictions on the outcome of any modification of the active site. Therefore, a targeted randomization of sets of amino acids in the active site coupled with a high-throughput screening of selected variants is considered to be the most promising strategy to influence the product formation of both enzymes. For the precise modification of interactions between substrate and protein we plan to incorporate functional groups that are not present in natural enzymes. For this, we will incorporate unnatural amino acids. The approach of Active Site Design is the targeted introduction of novel functional group coupled to a simultaneous variation of the molecular context. The optimal integration of the unnatural amino acids is aimed to achieve novel reactivities. Key for this proceeding is an interdisciplinary collaboration between enzyme engineering, organic synthesis and mechanistic studies.
StatusNot started
Effective start/end date1/09/2131/08/24