Primer utilization by DNA polymerase α-primase is influenced by its interaction with mcm10p Journal Article
| Authors: | Fien, K.; Cho, Y. S.; Lee, J. K.; Raychaudhuri, S.; Tappin, I.; Hurwitz, J. |
| Article Title: | Primer utilization by DNA polymerase α-primase is influenced by its interaction with mcm10p |
| Abstract: | Models of DNA replication in yeast and Xenopus suggest that Mcm10p is required to generate the pre-initiation complex as well as progression of the replication fork during the elongation of DNA chains. In this report, we show that the Schizosaccharomyces pombe Mcm10p/Cdc23p binds to the S. pombe DNA polymerase (pol) α-primase complex in vitro by interacting specifically with the catalytic p180 subunit and stimulates DNA synthesis catalyzed by the pol α-primase complex with various primed DNA templates. We investigated the mechanism by which Mcm10p activates the polymerase activity of the pol α-primase complex by generating truncated derivatives of the full-length 593-amino acid Mcm10p. Their ability to stimulate pol α polymerase activity and bind to single-stranded DNA and to pol α were compared. Concomitant with increased deletion of the N-terminal region (from amino acids 95 to 415), Mcm10p derivatives lost their ability to stimulate pol α polymerase activity and bind to single-stranded DNA. Truncated derivatives of Mcm10p containing amino acids 1-416 retained the pol α binding activity, whereas the C terminus, amino acids 496-593, did not. These results demonstrate that both the single-stranded DNA binding and the pol α binding properties of Mcm10p play important roles in the activation. In accord with these findings, Mcm10p facilitated the binding of pol α-primase complex to primed DNA and formed a stable complex with pol α-primase on primed templates. A mutant that failed to activate or bind to DNA and pol α, was not observed in this complex. We suggest that the interaction of Mcm10p with the pol α-primase complex, its binding to single-stranded DNA, and its activation of the polymerase complex together contribute to its role in the elongation phase of DNA replication. |
| Keywords: | controlled study; unclassified drug; nonhuman; dna polymerase; binding affinity; dna replication; protein function; protein analysis; cell cycle proteins; complex formation; enzyme inhibition; protein protein interaction; protein binding; enzyme activation; in vitro study; enzyme activity; dna; amino acid sequence; amino terminal sequence; binding energy; molecular interaction; yeast; catalysis; single stranded dna; enzyme kinetics; dna binding; fungal protein; schizosaccharomyces; schizosaccharomyces pombe proteins; complexation; schizosaccharomyces pombe; dna, fungal; derivatives; dna polymerase i; fungal dna; fungal enzyme; dna primase; protein mcm10; priority journal; article; pre-initiation complex; alpha primase; cell cycle protein 23 |
| Journal Title: | Journal of Biological Chemistry |
| Volume: | 279 |
| Issue: | 16 |
| ISSN: | 0021-9258 |
| Publisher: | American Society for Biochemistry and Molecular Biology |
| Date Published: | 2004-04-16 |
| Start Page: | 16144 |
| End Page: | 16153 |
| Language: | English |
| DOI: | 10.1074/jbc.M400142200 |
| PROVIDER: | scopus |
| PUBMED: | 14766746 |
| DOI/URL: | |
| Notes: | J. Biol. Chem. -- Cited By (since 1996):48 -- Export Date: 16 June 2014 -- CODEN: JBCHA -- Source: Scopus |
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