Optimization of the masking molecule for active-site-protected immobilization of Taq DNA polymerase and its application.

The method of oriented and activity-preserved immobilization of biologically active proteins based on concepts of active-site masking and kinetic control was further developed in this study. Minimal requirements for the masking DNA molecule were found to be a 5'overhang of 5-7 nucleotides and a double-stranded region of 11-13 bp to retain approximately 70% of the enzyme activity. The amplification range of protected immobilized (PIM) Taq DNA polymerase was over 1.2 kb. These data suggest that PIM Taq DNA polymerase can be used for various commercial applications.