Conformation of the origin of P1 plasmid replication : initiator protein induced wrapping and intrinsic unstacking

摘要: The origin of plasmid DNA replication in bacteriophage P1 has five 19 base-pair sites that bind the plasmid-encoded initiator, RepA. Here we show, using a band retardation assay, RepA can bend carries one or more binding sites. to supercoiled carrying sites, directly repeated and phased two turns B-DNA apart, absorbs about positive superhelical turn as determined by two-dimensional gel electrophoresis. This indicates is wrapped around consequence in-phase bending at individual RepA-DNA complexes did not show elevated sensitivity KMnO4, reagent specific for pyrimidine bases (T » C) unstacked DNA. wrapping RepA, therefore, does lead significant unwinding double helix. Extensive unwinding, suitable initiation replication, most likely requires participation factors other than RepA. We also noted thymine sequence 5′-ATC-3′, which there are 20 origin, all reacted KMnO4 strongly whether was present. preferential reactivity ATC sequences region, residues including those display fragment from pBR322. On strands selective supercoiling dependent. These results indicate includes bases, significance remains be determined.