Design of new enzymes for carbon-carbon bond formation

Project description

We are interested in the development of enantioselective proline-based biocatalysts for Michael-type addition reactions as a broadly useful C-C bond-forming methodology for the production of pharmaceuticals and fine chemicals. Using 4-oxalocrotonate tautomerase, which has a catalytic amino-terminal proline, we have recently developed protocols for enantioselective Michael-type reactions of a variety of aldehydes with various nitroolefins. These Michael-type reactions yield chiral building blocks for the synthesis of various GABA-B receptor agonists, which are important tools for neurobiology research.

The goal of the student project is to apply directed evolution to 4-oxalocrotonate tautomerase to increase the activity and scope of the enzyme in C-C bond formation. The student will start with the construction of a large library of 4-OT variants, using saturation mutagenesis at several combined positions in the substrate-binding pocket. Screening or selection will be used to obtain 4-OT variants with new are improved activities. The scope of nitroaldehyde products that can be synthesized using the newly designed enzymes will be explored by using UV spectroscopy, HPLC-MS, and NMR.

Organization

The master research project will be conducted in the Pharmaceutical Biotechnology group within the Dept. of Pharmaceutical Biology of Prof.dr. G.J. Poelarends. Our laboratory is well equipped for molecular biology, enzymology, protein engineering, and biocatalysis work. We offer a stimulating research environment for a student interested in these topics.

Requirements

We are looking for highly motivated students interested in performing state-of-the-art protein engineering work.

Contact

For further information you can contact Prof.dr. G.J. Poelarends, Dept. of Pharmaceutical Biology, GRIP, building 3215-room 9.21, Antonius Deusinglaan 1, 9713 AV GRONINGEN.