Double-strand cleavage and strand joining by the replication initiator protein of filamentous phage f1

D. Greenstein, K. Horiuchi

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The replication initiator protein (gene II protein (gpII)) of bacteriophage f1 is a multifunctional protein that plays central roles in initiation and termination of phage DNA replication. It introduces a nick at a specific site on the (+)-strand of supercoiled replicative form DNA. The 3'-hydroxyl end of the nick serves as the primer for (+)-strand rolling-circle replication. Upon completion of a round of synthesis, gpII cleaves and circularizes the displaced single strand. When Mn2+ is included in the buffer instead of Mg2+, gpII cleaves both strands. In this paper, we investigate the mechanism of the Mn2+-dependent double-strand cleavage activity of gpII. This reaction, unlike nicking in the presence of Mg2+, does not require superhelicity. The reaction proceeds in two kinetic steps: first nicking of the (+)-strand, and then cleavage of the (-)-strand. The nucleotide sequence requirement for nicking is reduced compared to that in the presence of Mg2+. The product of the double-strand cleavage has an unusual structure. The left end is a telomere-like hairpin since the (+)- and (-)-strands are joined, as demonstrated by base sequencing. The right end has a one-base 3'-overhang. This reaction probably reflects the cleavage-joining activity of gpII in the termination event.

Original languageEnglish (US)
Pages (from-to)12627-12632
Number of pages6
JournalJournal of Biological Chemistry
Issue number21
StatePublished - 1989


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