Biochemical Characterization of the Staphylococcus aureus PcrA Helicase and Its Role in Plasmid Rolling Circle Replication
| Autor: | Tseh-Ling Chang, Asma Naqvi, Rajan Munshi, Syam P. Anand, M. Gabriela Kramer, Saleem A. Khan |
|---|---|
| Rok vydání: | 2002 |
| Předmět: |
DNA Replication
DNA Bacterial Staphylococcus aureus Molecular Sequence Data Origin of replication Biochemistry chemistry.chemical_compound Plasmid Bacterial Proteins Molecular Biology DNA Primers Base Sequence biology DNA Helicases Helicase Cell Biology PcrA RNA Helicase A Molecular biology Cell biology chemistry Rolling circle replication biology.protein Replisome DNA Plasmids Protein Binding |
| Zdroj: | Journal of Biological Chemistry. 277:45880-45886 |
| ISSN: | 0021-9258 |
| Popis: | Previous genetic studies have suggested that a putative chromosome-encoded helicase, PcrA, is required for the rolling circle replication of plasmid pT181 in Staphylococcus aureus. We have overexpressed and purified the staphylococcal PcrA protein and studied its biochemical properties in vitro. Purified PcrA helicase supported the in vitro replication of plasmid pT181. It had ATPase activity that was stimulated in the presence of single-stranded DNA. Unlike many replicative helicases, PcrA was highly active as a 5' --3' helicase and had a weaker 3' --5' helicase activity. The RepC initiator protein encoded by pT181 nicks at the origin of replication and becomes covalently attached to the 5' end of the DNA. The 3' OH end at the nick then serves as a primer for displacement synthesis. PcrA helicase showed an origin-specific unwinding activity with supercoiled plasmid pT181 DNA that had been nicked at the origin by RepC. We also provide direct evidence for a protein-protein interaction between PcrA and RepC proteins. Our results are consistent with a model in which the PcrA helicase is targeted to the pT181 origin through a protein-protein interaction with RepC and facilitates the movement of the replisome by initiating unwinding from the RepC-generated nick. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|