Enzyme immobilization on hydroxyapatite: a case study

Enzymes are efficient biocatalysts which operate under mild conditions with high selectivity, low toxicity and negligible by-product. These characteristics make them important tools in making chemical processes greener. However, enzymes are difficult to recover and reuse after the reaction and their activity can be lost due to adverse process conditions. A common strategy to solve these problems is the immobilization of the enzyme on a solid support. Hydroxyapatite (HAP) is a biocompatible inorganic material which appears to be feasible for this task thanks to its structural stability, non-toxicity, large surface area and ease of surface modification. Moreover, it can be obtained from some waste (e.g. ashes from waste-to-energy plants, the fish supply chain, the avian supply chain, etc.) in agreement with circular economy principles. As a first approach by our research group, gamma-glutamyl transferase from Bacillus subtilis (BsGGT) was chosen as a model enzyme to study the immobilization process on HAP. HAP was synthesised according to a literature-reported procedure. Enzyme immobilization was initially attempted exploiting the electrostatic interactions between the charged groups on the surfaces of both HAP and the enzyme, simple by mixing an enzyme solution and a hydroxyapatite suspension under controlled conditions (pH, temperature). Different particle sizes and experimental set-up were investigated and the supported BsGGT was tested as a biocatalyst in the gamma-glutamylation of L-methionine as a model reaction. In the meantime, hydroxyapatite functionalization was studied to achieve a covalent enzyme immobilization.