L-lysine-coated magnetite nanoparticles (L-lys@Fe3O4) were employed for the magnetic immobilization of recombinant Escherichia coli cells producing extracellular L-asparaginase II. Impact of magnetic immobilization on the plasmid instability and development of a plasmid-free cell community was investigated. Also, undesirable interactions of nanoparticles with the produced enzyme leading to enzyme inactivation or enzyme entrapment were evaluated. Employed L-lys@Fe3O4 nanoparticles were ~ 12.5–22 nm in size with 42 emu/g saturation magnetization. Immobilized cells were magnetically sensitive and completely harvested by applying an external magnetic field (immobilization efficiency equal to 100%). L-asparaginase activity analysis determined that L-lys@Fe3O4 nanoparticles have no inactivation effect on the produced recombinant L-asparaginase II. But a little fraction of the enzyme can be expected to be entrapped within the nanoparticles. However, in contrast to total amount of enzyme the entrapped enzyme was negligible. Due to magnetic immobilization, a small (4.3%) reduction in the community of plasmid-bearing cells was observed but this value was not statistically significant. These findings can pave the way for further applications of magnetic immobilization to immobilize recombinant bacterial cells.