Construction and Catalytic Study of Affinity Peptide Orientation and Light Crosslinking Immobilized Sucrose Isomerase

A novel affinity peptide orientation and light-controlled covalent immobilized method was developed. Sucrose isomerase (SI) was selected as the model enzyme. Molecular simulation was first performed to select the targeted immobilization region. Subsequently, a short peptide (H2N-VNIGGX-COOH, VG) with high affinity to this region was rationally designed. Thereafter, 4-benzoyl-l-phenylalanine with the photosensitive group of benzophenone was introduced. Then, the affinity between the ligand and the SI was validated using molecular dynamics simulation. Thereafter, the SI was directionally immobilized onto the surface of the epoxy resin (EP) guided by VG via photo-crosslinking, and thus the oriented photo-crosslinking enzymes were obtained. The enzymatic activity, thermostability, and reusability of the affinity directional photo-crosslinked immobilized sucrose isomerase (hv-EP-VG-SI) were systematically studied. The oriented immobilization enzymes were significantly improved in recycling and heat resistance. Moreover, hv-EP-VG-SI retained more than 90% of the original activity and 50% of the activity after 11 cycles.

Automated summary

A novel method of affinity peptide orientation and light-controlled covalent immobilization was developed in this study. The targeted immobilization region was selected using molecular simulation, and a short peptide with high affinity was designed and validated. Through directional photo-crosslinking, the enzymes were immobilized onto the surface of the epoxy resin, resulting in improved recycling and heat resistance.