Biosynthesis of gold cluster nanozyme within the structure of TetX2 monooxygenase Protein

The synthesis of gold nanoclusters within protein structures, such as TetX2 monooxygenase, presents a promising approach for obtaining nanostructures with enzyme-like activity. This is attributed to the presence of various amino acid groups that serve as biological templates for the formation and reduction of gold. The low toxicity, biocompatibility, solubility in aqueous systems, and specific fluorescence spectrum of these nanostructures enhance their utility in the detection of various analytes. The research involved expressing and purifying the recombinant enzyme TetX2, synthesizing gold on the TetX2 substrate, and investigating the effect of nanoclusters on the activity and structure of the TetX2 enzyme. The synthesis of gold nanoclusters(AuNCs) was carried out within the structure of TetX2. Finally, the accuracy of the AuNCs synthesis was verified by examining the morphology, size, spectroscopy, and catalytic activity. Results showed changes in the tertiary structure of the TetX2 enzyme in the presence of HAuCl4, loss of enzyme activity after the formation of gold nanoclusters, and successful incorporation of gold into the structure of TetX2. TetX2@AuNCs emitted blue light at 450 nm, and their catalytic properties were demonstrated through color changes in a chromogenic substrate (3,3',5,5'-Tetramethylbenzidine-H2O2), indicating their authenticity. TetX2 is proposed as a new bio-template forthe synthesis of gold nanoclusters, with potential applications in biosensor design (Graphic abstract).