Recombinant Escherichia coli systems expressing organophosphorous hydrolase (OPH) have been used for detoxifying toxic organophosphate compounds. However, a whole cell biocatalyst system has an intrinsic problem due to substrate diffusion limitation by its cell membrane. As a strategy for reducing this diffusion barrier limitation to enhance whole cell biocatalytic activity, we engineered E. coli cells to target OPH on cell surface using ice nucleation protein (InaK) as a surface targeting motif, especially N-terminal domain of InaK (InaK-N). The whole cell OPH activities of the cells expressing InaK/OPH fusion constructs were higher (~2.5-fold for InaK-N and ~5.7-fold for combined N- and C-terminal domain of InaK (InaK-NC)) than that of the cells expressing cytosolic OPH. Interestingly, the membrane targeting efficiency of the cells expressing InaK-N/OPH fusion proteins was ~2.2-fold higher compared to the cells expressing InaK-NC/OPH even though both whole cell and total cell lysate OPH activities were lower. Therefore, we found that the small size N-terminal domain of InaK is more efficient for targeting OPH on the cell surface, and the surface display of OPH using N-terminal InaK domain can reduce the mass-transfer problem in whole cell bioconversion system.