Interactions of human MCM2-7 proteins with the proteins of TIM, TIPIN, an amino-terminal fragment of Rb, and p27 were examined by co-immuno-precipitation experiment using cell lysates of co-expressed insect cells. TIM and TIPIN, both of which are involved in regulation of DNA replication fork progression, mainly interacted with MCM3-7 proteins. The amino-terminal fragment of Rb, which inhibits DNA replication in Xenopus egg extracts, was able to bind with MCM3 and MCM6 proteins in addition to MCM7 protein. In contrast, p27 was not able to bind any MCM2-7 proteins under the comparable conditions. These results indicate that the proteins, which are known to interact with MCM proteins, bind with MCM2-7 proteins with different affinities and specificities.