Cholesterol oxidase(ChOx) enzyme isolated from Pseudomonas aeruginosa PseA(ChOxP) and Rhodococcus erythropolis MTCC 3951(ChOxR) strains as well as a commercial variant produced by Streptomyces sp.(ChOxS) were immobilized on silane modified iron(II, III) oxide magnetic nanoparticles(MNP) by covalent coupling methods. The nanobiocatalysts in case of ChOxP, ChOxR and ChOxS, retained 71, 91 and 86% of cholesterol oxidase activity respectively, as compared to their soluble counterparts. The catalytic efficiency of the immobilized enzymes on nanoparticles was more than 2.0 times higher than the free enzyme. They also showed enhanced pH and thermal stability. After 10 cycles of operation, the MNP-bioconjugates retained 50, 52 and 51% of residual activity in case of ChOxP, ChOxR and ChOxS respectively. The presence of enzyme on nanoparticles was confirmed by FTIR, SEM and TEM. The nanobiocatalysts were used for the biotransformation of cholesterol and 7-ketocholesterol to 4-cholesten-3-one and 4-cholesten-3, 7-dione respectively, which are industrially and medically important steroid precursors.