Immobilization of yeast alcohol dehydrogenase on magnetic nanoparticles for improving its stability

Yeast alcohol dehydrogenase (YADH) was immobilized covalently on Fe3O4 magn etic nanoparticles (10.6 nm) via carbodiimide activation. The immobilizatio n process did not affect the size and structure of magnetic nanoparticles. The YADH-immobilized magnetic nanoparticles were superparamagnetic with a s aturation magnetization of 61 emu g(-1), only slightly lower than that of t he naked ones (63 emu g(-1)). Compared to the free enzyme, the immobilized YADH retained 62% activity and showed a 10-fold increased stability and a 2 .7-fold increased activity at pH 5. For the reduction of 2-butanone by immo bilized YADH, the activation energies within 25-45 degreesC, the maximum sp ecific activity, and the Michaelis constants for NADH and 2-butanone were 2 7 J mol(-1), 0.23 mol min(-1) mg(-1), 0.62 mM, and 0.43 M, respectively. Th ese results indicated a structural change of YADH with a decrease in affini ty for NADH and 2-butanone after immobilization compared to the free enzyme .