Processive DNA helicase activity of the minichromosome maintenance proteins 4, 6, and 7 complex requires forked DNA structures.
The minichromosome maintenance (Mcm) proteins 2-7 are required for both the initiation and elongation steps of chromosomal DNA replication. Previous studies have shown that the Mcm complex consisting of the Mcm 4, 6, and 7 proteins contains 3' to 5' DNA helicase activity with limited processivity (displacing duplex DNA regions up to 30 nt). In this report, we show that the presence of both 5' and 3' single-stranded tails in DNA helicase substrates is essential for the processive helicase activity of the Mcm complex. The presence of both 5' and 3' tails facilitated the formation of double heterohexameric complexes of Mcm4/6/7 on substrate DNA, which appeared to be essential for the processive helicase activity. The double heterohexameric complex of Mcm4/6/7, in the presence of a single-strand DNA binding protein, is capable of unwinding duplex DNA region of about 600 bp in length. These results support the hypothesis that the Mcm4/6/7 complex can function as a replication helicase.
Lee
J K
JK
Graduate Program in Molecular Biology, Memorial Sloan-Kettering Cancer Center, New York, NY 10021, USA.
Hurwitz
J
J
D018797:Cell Cycle Proteins / Q000378:metabolism*; D002499:Centrifugation, Density Gradient; D002868:Chromosomal Proteins, Non-Histone; D003432:Cross-Linking Reagents / Q000378:metabolism; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004265:DNA Helicases / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004587:Electrophoresis, Agar Gel; D004789:Enzyme Activation; D046911:Macromolecular Substances; D064148:Minichromosome Maintenance Complex Component 4; D064151:Minichromosome Maintenance Complex Component 6; D064168:Minichromosome Maintenance Complex Component 7; D009687:Nuclear Proteins / Q000378:metabolism*; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D011994:Recombinant Proteins / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D012568:Schizosaccharomyces / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity
D018797:Cell Cycle Proteins; D002868:Chromosomal Proteins, Non-Histone; D003432:Cross-Linking Reagents; D004268:DNA-Binding Proteins; D046911:Macromolecular Substances; D009687:Nuclear Proteins; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D004247:DNA; D004265:DNA Helicases; C092802:MCM4 protein, S cerevisiae; C491207:MCM6 protein, S cerevisiae; C491088:MCM7 protein, S cerevisiae; D064148:Minichromosome Maintenance Complex Component 4; D064151:Minichromosome Maintenance Complex Component 6; D064168:Minichromosome Maintenance Complex Component 7