Characterization of the replication initiator Orc1/Cdc6 from the Archaeon Picrophilus torridus
| Autor: | Jasmine Arora, Kasturi Goswami, Swati Saha |
|---|---|
| Rok vydání: | 2013 |
| Předmět: |
DNA Replication
Binding Sites Picrophilus torridus Archaeal Proteins DNA replication Origin Recognition Complex Computational Biology Eukaryotic DNA replication Thermoplasmales Articles Biology Pre-replication complex biology.organism_classification Microbiology DnaA Replication factor C DNA Archaeal Biochemistry Proliferating Cell Nuclear Antigen Origin recognition complex Protein Multimerization Molecular Biology Picrophilus Protein Binding |
| Zdroj: | Journal of bacteriology. 196(2) |
| ISSN: | 1098-5530 |
| Popis: | Eukaryotic DNA replication is preceded by the assembly of prereplication complexes (pre-RCs) at or very near origins in G1 phase, which licenses origin firing in S phase. The archaeal DNA replication machinery broadly resembles the eukaryal apparatus, though simpler in form. The eukaryotic replication initiator origin recognition complex (ORC), which serially recruits Cdc6 and other pre-RC proteins, comprises six components, Orc1-6. In archaea, a single gene encodes a protein similar to both the eukaryotic Cdc6 and the Orc1 subunit of the eukaryotic ORC, with most archaea possessing one to three Orc1/Cdc6 orthologs. Genome sequence analysis of the extreme acidophile Picrophilus torridus revealed a single Orc1/Cdc6 (PtOrc1/Cdc6). Biochemical analyses show MBP-tagged PtOrc1/Cdc6 to preferentially bind ORB (origin recognition box) sequences. The protein hydrolyzes ATP in a DNA-independent manner, though DNA inhibits MBP-PtOrc1/Cdc6-mediated ATP hydrolysis. PtOrc1/Cdc6 exists in stable complex with PCNA in Picrophilus extracts, and MBP-PtOrc1/Cdc6 interacts directly with PCNA through a PIP box near its C terminus. Furthermore, PCNA stimulates MBP-PtOrc1/Cdc6-mediated ATP hydrolysis in a DNA-dependent manner. This is the first study reporting a direct interaction between Orc1/Cdc6 and PCNA in archaea. The bacterial initiator DnaA is converted from an active to an inactive form by ATP hydrolysis, a process greatly facilitated by the bacterial ortholog of PCNA, the β subunit of Pol III. The stimulation of PtOrc1/Cdc6-mediated ATP hydrolysis by PCNA and the conservation of PCNA-interacting protein motifs in several archaeal PCNAs suggest the possibility of a similar mechanism of regulation existing in archaea. This mechanism may involve other yet to be identified archaeal proteins. |
| Databáze: | OpenAIRE |
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