Figure 4

Oligomeric states and interactions of N-terminal fragments of Mcm4, 6 and 7. (A) Gel filtration chromatography profiles of N-terminal fragments of Mcm4, 6 and 7. Schematic of each fragment was shown in accordance with its gel filtration profile. N-terminal fragments of Mcm6 were aligned with the zinc finger motif and a 62 amino acid residues protruding N-termini was shown. 7N1a, separated monomeric 7 N1 fragment; 7N1b, separated dimeric 7 N1 fragment. Gel filtration analysis was carried out a described under “Materials and Methods”. (B) In vitro incubation of purified N-terminal fragments of Mcm4, 6 and 7. Interactions among the N-terminal fragments of Mcm4, 6 and 7 were characterized by gel filtration analysis. Samples from peak fractions (pointed by arrows) were quantitated by SDS-PAGE and mixed together in approximate equal molar ratio. The mixture was buffer-exchanged to 50 mM NaCl, 50 mM Tris pH8 and 1 mM DTT and then incubated on ice for 30 minutes. For 7 N1 and 7 N2, only samples from peak fraction of monomeric states were used. The incubation mixtures were subjected to gel filtration analysis and no large complex was detected. Two groups of N-terminal fragments of Mcm4, 6 and 7 were used, as shown in top and bottom panels.