Roles of Mcm7 and Mcm4 subunits in the DNA helicase activity of the mouse Mcm4/6/7 complex.
Mcm, which is composed of six structurally related subunits (Mcm2-7), is essential for eukaryotic DNA replication. A subassembly of Mcm, the Mcm4/6/7 double-trimeric complex, possesses DNA helicase activity, and it has been proposed that Mcm may function as a replicative helicase at replication forks. We show here that conserved ATPase motifs of Mcm7 are essential for ATPase and DNA helicase activities of the Mcm4/6/7 complex. Because uncomplexed Mcm7 displayed neither ATPase nor DNA helicase activity, Mcm7 contributes to the DNA helicase activity of the Mcm complex through interaction with other subunits. In contrast, the Mcm4/6/7 complex containing a zinc finger mutant of Mcm4 with partially impaired DNA binding activity exhibited elevated DNA helicase activity. The Mcm4/6/7 complex containing this Mcm4 mutant tended to dissociate into trimeric complexes, suggesting that the zinc finger of Mcm4 is involved in subunit interactions of trimers. The Mcm4 mutants lacking the N-terminal 35 or 112 amino acids could form hexameric Mcm4/6/7 complexes, but displayed very little DNA helicase activity. In conjunction with the previously reported essential role of Mcm6 in ATP binding (You, Z., Komamura, Y., and Ishimi, Y. (1999) Mol. Cell. Biol. 19, 8003-8015), our data indicate distinct roles of Mcm4, Mcm6, and Mcm7 subunits in activation of the DNA helicase activity of the Mcm4/6/7 complex.
You
Zhiying
Z
Department of Cell Biology, Tokyo Metropolitan Institute of Medical Science, 18-22 Honkomagome 3-chome, Bunkyo-ku, Tokyo 113-8613, Japan. zyzhiying@rinshoken.or.jp
Ishimi
Yukio
Y
Masai
Hisao
H
Hanaoka
Fumio
F
D000251:Adenosine Triphosphatases / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D017124:Conserved Sequence; D004265:DNA Helicases / Q000737:chemistry / Q000378:metabolism*; D017931:DNA Primers; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D051379:Mice; D064148:Minichromosome Maintenance Complex Component 4; D064151:Minichromosome Maintenance Complex Component 6; D064168:Minichromosome Maintenance Complex Component 7; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism*; D016335:Zinc Fingers
D018797:Cell Cycle Proteins; D017931:DNA Primers; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D021122:Protein Subunits; D000251:Adenosine Triphosphatases; D004265:DNA Helicases; C099243:MCM4 protein, human; C491205:Mcm6 protein, mouse; C491086:Mcm7 protein, mouse; D064148:Minichromosome Maintenance Complex Component 4; D064151:Minichromosome Maintenance Complex Component 6; D064168:Minichromosome Maintenance Complex Component 7