Escherichia coli SeqA protein affects DNA topology and inhibits open complex formation at oriC
Chromosome replication in Escherichia coli is initiated by the DnaA protein. Binding of DnaA to the origin, oriC , followed by formation of an open complex are the first steps in the initiation process. Based on in vivo studies the SeqA protein has been suggested to function negatively in the initia...
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Veröffentlicht in: | The EMBO journal 1999-09, Vol.18 (17), p.4882-4888 |
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Sprache: | eng |
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Zusammenfassung: | Chromosome replication in
Escherichia coli
is initiated by the DnaA protein. Binding of DnaA to the origin,
oriC
, followed by formation of an open complex are the first steps in the initiation process. Based on
in vivo
studies the SeqA protein has been suggested to function negatively in the initiation of replication, possibly by inhibiting open complex formation.
In vitro
studies have shown that SeqA inhibits
oriC
‐dependent replication. Here we show by KMnO
4
probing that SeqA inhibits open complex formation. The inhibition was not caused by prevention of DnaA binding to the
oriC
plasmids, indicating that SeqA prevented strand separation in
oriC
either directly, by interacting with the AT‐rich region, or indirectly, by changing the topology of the
oriC
plasmids. SeqA was found to restrain the negative supercoils of the
oriC
plasmid. In comparison with the effect of HU on plasmid topology, SeqA seemed to act more cooperatively. It is likely that the inhibition of open complex formation is caused by the effect of SeqA on the topology of the plasmids. SeqA also restrained the negative supercoils of unmethylated
oriC
plasmids, which do not bind SeqA specifically, suggesting that the effect on topology is not dependent on binding of SeqA to a specific sequence in
oriC
. |
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ISSN: | 0261-4189 1460-2075 1460-2075 |
DOI: | 10.1093/emboj/18.17.4882 |