ProjectsDesign of WW-domain based mini enzymes

The biological catalysis is impressively efficient and is characterized by high substrate specificity as well as stereo and regioselectivity. However, enzymes are usually huge molecules with a complex three-dimensional structure and an insufficiently understood sequence-structure-function relationship. Therefore, small peptides with predictable and reliable folding properties, mainly coiled-coil and β-amyloid scaffolds, have been elucidated as platforms for the design of mini-enzymes that mimic the biological activity of enzymes but also catalyse reactions that do not occur in nature.

We are interested in converting the protein-binding WW domain into catalytically active peptide scaffolds. In this context we pursue different strategies: For example, we use a split WW domain approach to combinatorially identify functional scaffolds. For this purpose, we split the three-stranded β-motif in the first loop and reconstitute the fragmented peptide by coiled-coil association. This method allows us to mix and combine several variants of WW domain fragments, thus increasing the number of possible WW domains to be tested by n2 (n = number of WW-domain variants).

A second approach comprises the top-down engineering of native WW domains. For example, we engineered a His3-site on the β-sheet surface to create a Zinc-coordination site similar to the active center of the human carbonic anhydrase II. This WW domain, WW-CA, shows highly reversible on-off-folding when Zn2+ is added and removed. Additionally, the peptide is able to catalyze the hydrolysis of p-NPA. Although the catalytic activity of WW-CA is lower than that of native hydrolases, this pioneering project clearly demonstrates the potential of WW domains as scaffolds for the design of biomimetic catalysts. Since WW-CA also binds to Cu2+ and Cu+, we are confident that catalysis is not limited to hydrolysis but can be extended to redox reactions.

Design of Mini-Enzymes

Project Lead

Jun.-Prof. Dr. Franziska Thomas
Organic Chemistry