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Volume 271, Number 48, Issue of November 29, 1996 pp. 31000-31007
©1996 by The American Society for Biochemistry and Molecular Biology, Inc.

Activation of Replication Origins in phi 29-related Phages Requires the Recognition of Initiation Proteins to Specific Nucleoprotein Complexes

(Received for publication, July 18, 1996)

Raimundo Freire , Manuel Serrano , Margarita Salas and José M. Hermoso

From the Centro de Biología Molecular "Severo Ochoa" (CSIC-UAM), Universidad Autónoma, Cantoblanco, 28049 Madrid, Spain

Protein p6 of Bacillus subtilis phage phi 29 activates the initiation of viral DNA replication by forming a multimeric nucleoprotein complex at the origins of replication, located at both ends of the linear genome. This activation requires a precise positioning of the protein p6 array with respect to the initiation site. To investigate this activation mechanism, we have purified the phi 29 protein p6 counterparts from the related phages Nf and GA-1 and analyzed the formation of complexes with DNA. In the homologous protein p6-DNA complexes the phi 29 and Nf protein arrays showed an identical positioning, different than that of the GA-1 protein array. In contrast, in the heterologous complexes the protein showed a different arrangement except in the case of the Nf protein-phi 29 DNA complex. We have also purified the proteins involved in the initiation of replication (terminal protein and DNA polymerase) from phages Nf and GA-1 and measured the ability of the different p6 proteins to activate homologous and heterologous replication origins. The results obtained indicate that the activation requires not only the formation of a specific nucleoprotein complex but also its specific recognition by the proteins involved in the initiation of DNA replication.


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