The main objective of research is to understand the structural basis of metalloprotein functions, with particular interest in iron and copper-containing enzymes. The proteins are purified from various organisms, the corresponding genes identified, cloned, sequenced, and the proteins finally heterologously overexpressed (Escherichia coli, Pichia pastoris, Chinese hamster ovary-, HEK-cell lines).
Exchange of amino acids by either site-directed or random mutagenesis in combination with biophysical methods (electron paramagnetic resonance spectroscopy, resonance Raman spectroscopy, UV-Vis- and fluorescence spectroscopy, circular dichroism spectroscopy, differential scanning calorimetry), transient- and steady-state kinetic investigations (multi-mixing stopped-flow spectroscopy) and X-ray crystallography allow elucidation of structure and function including detailed characterization of the corresponding redox intermediates, which are relevant in catalysis. In addition we ask for the role of post-translational modifications in the prosthetic groups and the protein matrix in catalysis of oxidoreductases.
We also prepare (recombinant) enzymes for customers (http://www.myeloperoxidase.com/) and provide quality control and kinetic analyses of (recombinant) proteins.
In addition, we are also active in the protein engineering field. In these projects, we design proteins with new or improved functions by directed evolution. These protein engineering methods can be used for numerous applications, such as the development of novel strategies in cancer immunotherapy or new components for diagnostic assays. Furthermore, we also use directed evolution to simulate resistance mechanisms in cancer cells.