C-terminal regions of the protein phosphatases PP1 and PP2B were seldom studied. C-terminal 24 amino acids of PP1 was deleted, its enzymatic activity increased 3-fold while its stability declined. When the truncated PP1 was fused with the terminal (residues 483–511) of PP2B, both its enzymatic activity and its stability remained low. This indicates that the termini of PP2B and PP1 have inhibitory effect on the catalytic domain of PP1. PP1-(1–306) and PP1wt differ in their activation by metal ions, showing that the sites interacting with metal ions are not located in its C-terminus; while metal ions activated notably to PP1/PP2B chimera. In addition, the sensitivity results of PP1-(1–306) to the inhibitors, TM and NCTD, proved that these two inhibitors also did not bind to the C-terminus. However, the IC50s of PP1/PP2B chimera were higher than for PP1-(1–306), indicating that the C-terminal region interferes interactions with these inhibitors to some extent. Although 483–511 segment of PP2B was not the functional domain, it played important role in interaction with metal ions and inhibitors. It further indicates although PP1 and PP2B have high sequence identity, their non-conserved termini have different roles.