Immobilized enzymes have provided tremendous advantages for the efficient production of biomaterials. There is increasing demand on simple and convenient protein immobilization methods because protein microarray is emerging as a cutting-edge technology for the Proteome analysis and diagnosis. It has been shown that a poly-lysine tag facilitates protein purification and refolding processes. This study demonstrates that the same poly-lysine tag can be employed for the immobilization of enzyme on a solid support without deterioration of its enzymatic characteristics. Cyclodextrin glycosyltransferase (CGTase) derived from Bacillus macerans was fused to consecutive 10 lysine residues (CGTK10ase) and electrostatically immobilized on a cation exchanger. Analyses on the binding characteristics, effects of pH and temperature on enzyme stability and operational stability indicate that the poly-lysine tag is also effective for non-covalent immobilization of CGTase. Though the poly-lysine-mediated immobilization is reversible, binding force is strong enough to block protein leakage from the solid support at neutral and basic pH. (C) 2004 Elsevier Inc. All rights reserved.