FIGURE 2.

An N-terminal Mcm2 region mediates interaction with Dbf4. A, Mcm2WT and mutant cassettes used in this study. The location of the conserved MCM box, including Walker A, Walker B, and arginine finger motifs, is indicated. B, C, and E, two-hybrid assays carried out using bait construct pEG-Dbf4. pJG-Mcm2 (WT), -Mcm2(1–504), -Mcm2(505–868), -Mcm2Δ63, and -Mcm2Δ2–4,10–63 were used as prey. The average of three replicates is shown ± S.D. (error bars). Immunoblot analyses to verify bait and prey expression were carried out as described for Fig. 1. D and F, immunoprecipitation (IP) of Myc-tagged Dbf4. Shown are immunoblots of IP and supernatant (S) fractions detected with monoclonal anti-HA (Mcm2 detection) and anti-Myc antibodies (Dbf4 detection). 20 μg of input and one-fourth of the final bead suspension were loaded.