Structure-Based Modeling of the Mechanical Behavior of Cross-Linked Enzyme Crystals
Because of their high volumetric catalytic activity, in addition to their high chemical and thermal resistances, enzymes in the form of protein crystals are an excellent choice for application as immobilized biocatalysts. However, mechanical stability is a requirement for the processability of immobilisates, in addition to the protein crystals retaining their enzymatic activity, and this is closely related to the crystal structure. In this study, the influence of protein engineering on the mechanical stability of cross-linked enzyme crystals (CLECs) was investigated using a genetically modified model protein in which additionally cysteines were introduced on the protein surface for targeted cross-linking. The results showed that the mechanical stability of crystals of the mutant proteins in the native form was decreased compared to native wild-type crystals. However, specific cross-linking of the introduced amino acid residues in the mutant proteins resulted in their increased mechanical stability compared to wild-type CLECs. In order to determine the correlation between the crystal structure and the resulting mechanical properties of CLECs to enable targeted cross-linking, a previously developed model was revised and then used for the two model proteins. This model can explain the mechanically investigated relationships, such as the anisotropic crystal behavior and the influence of a linker or mutation on the micromechanical properties and, hence, can be helpful for the tailor-made production of CLECs.