Pillared interlayered clays (PILCs) was a versatile kind of nanoporous material, however, no research on protein immobilized on PILCs had been reported. The objective of the paper was to study application of PILCs in enzyme immobilization. [Methods] Horseradish Peroxidase (HRP) used as model protein was to be bounded on Al-PILC. The support and immobilized enzyme were characterized by way of N₂-adsorption and desorption, XRD and FTIR. The conditions of immobilization such as dose of support, pH value and immobilization time were investigate by comparison of enzyme immobilization yield and catalytic performance of immobilized enzyme in the treatment of phenolic wastewater. And kinetic properties of immobilized and free HRP were also studied through a series of catalytic reactions. [Results] HRP could be successfully attached to Al-PILC on simple conditions, and the optimum immobilization conditions were ascertained. HRP immobilized on Al-PILC exhibited well catalytic performance. The Km values of immobilized HRP were higher than the corresponding values obtained for soluble enzyme. [Conclusions] Al-PILC was a promising material used as the support for enzyme immobilization. The enzymatic reaction mechanism were unaffected by immobilization.