Covalent immobilization of Bacillus subtilis lipase A on Fe_3O_4 nanoparticles by aldehyde tag: An ideal immobilization with minimal chemical modification

摘要

Lipase A from Bacillus subtilis with modified aldehyde tags was successfully immobilized covalently on Fe3O4 monodispersed magnetite microspheres and greatly improves thermal stability. The properties of immobilized enzyme were investigated. Commercial magnetic particles embedded in SiO2 nanoparticles with size of 100 nm were amino-functionalised. Bacilhis subtilis lipase A (BsLA) mutant contained eight changed amino acid, which improved the solubility of the enzyme and reduced the possibility of enzyme coagulation. BsLA 8M was constructed with LCTPSR tag to introduce an aldehyde group. The magnetic particles were covalently bonded BsLA 8M by Schiff Base reaction. The immobilization efficiency was over 90%, and the activity of the immobilized lipase was enhanced. The optimum immobilization conditions were phosphate buffer pH 8.5, carrier concentration of 500 mg/g and incubated and shaken for 6 h at 25 degrees C. The optimum pH and temperature of the immobilized enzyme was 8.0 and 70 degrees C, respectively. The activity of the immobilized lipase remained at 90% after 10 cycles. The construction of BsLA 8M with aldehyde tags and optimization of immobilization conditions resulted in enhanced thermal stability, easier recycling and higher optimum reaction temperature compared with those of free lipase.