Hard X-ray Fluorescence mapping to decipher the deactivation processes of immobilised metallo-enzyme systems

Immobilisation of enzymatic cascades in porous materials is ideal to perform multi-enzymatic biotransformations in industrial settings. Yet, they suffer from exhaustion processes that hampers their use for long operational periods. The exact mechanism of inactivation is unknown until now, but spatial re-organisation of enzymes across the material or loss of metal ions from their active centers seems to be relevant. Unfortunately, optical microscopy is not ideal to study those events, as labelling enzymes with fluorescent tags can alter their catalytic properties. We intend to acquire X-Ray Fluorescence maps at ID16A on methacrylate beads carrying a cascade formed by Zn and Cu-dependent enzymes used to transform diols to relevant lactones, both before and after their operational run. This will provide valuable information on the importance of enzymatic re-organisation and metal leaching for the deactivation of immobilised multi-enzyme systems upon operation in chemical manufacturing.