Yeast alcohol dehydrogenase (YADH) was immobilized covalently on Fe3O4 magnetic nanoparticles (10.6 nm) via carbodiimide activation. The immobilization process did not affect the size and structure of magnetic nanoparticles. The YADH-immobilized magnetic nanoparticles were superparamagnetic with a saturation magnetization of 61 emu g(-1), only slightly lower than that of the 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 immobilized YADH, the activation energies within 25-45 degreesC, the maximum specific activity, and the Michaelis constants for NADH and 2-butanone were 27 J mol(-1), 0.23 mol min(-1) mg(-1), 0.62 mM, and 0.43 M, respectively. These results indicated a structural change of YADH with a decrease in affinity for NADH and 2-butanone after immobilization compared to the free enzyme.