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Interactions between the RepB initiator protein of plasmid pMV158 and two distant DNA regions within the origin of replication.

Authors :
Ruiz-Masó JA
Lurz R
Espinosa M
del Solar G
Source :
Nucleic acids research [Nucleic Acids Res] 2007; Vol. 35 (4), pp. 1230-44. Date of Electronic Publication: 2007 Jan 31.
Publication Year :
2007

Abstract

Plasmids replicating by the rolling circle mode usually possess a single site for binding of the initiator protein at the origin of replication. The origin of pMV158 is different in that it possesses two distant binding regions for the initiator RepB. One region was located close to the site where RepB introduces the replication-initiating nick, within the nic locus; the other, the bind locus, is 84 bp downstream from the nick site. Binding of RepB to the bind locus was of higher affinity and stability than to the nic locus. Contacts of RepB with the bind and nic loci were determined through high-resolution footprinting. Upon binding of RepB, the DNA of the bind locus follows a winding path in its contact with the protein, resulting in local distortion and bending of the double-helix. On supercoiled DNA, simultaneous interaction of RepB with both loci favoured extrusion of the hairpin structure harbouring the nick site while causing a strong DNA distortion around the bind locus. This suggests interplay between the two RepB binding sites, which could facilitate loading of the initiator protein to the nic locus and the acquisition of the appropriate configuration of the supercoiled DNA substrate.

Details

Language :
English
ISSN :
1362-4962
Volume :
35
Issue :
4
Database :
MEDLINE
Journal :
Nucleic acids research
Publication Type :
Academic Journal
Accession number :
17267412
Full Text :
https://doi.org/10.1093/nar/gkl1099