Description:
<jats:title>Summary</jats:title><jats:p>We analysed the enzymatic activity (strand dis‐placement) of the <jats:italic>Escherichia coli</jats:italic> DnaB helicase on a mirror‐image pair of oligonucleotide‐based substrates mimicking the unwound replication origin <jats:italic>oriC</jats:italic>. Loading of the helicase complex occurred exclusively to the single‐stranded ‘lower strand’ part of the substrates. Full helicase activity required DnaA bound to the double‐stranded part of the substrates (<jats:italic>oriC</jats:italic> DnaA box R1) and to their single‐stranded ‘upper strand’ part. We assume that <jats:italic>in vivo</jats:italic> DnaA also loads the first of two helicase complexes – required for the assembly of two replication forks – to the lower strand of <jats:italic>oriC</jats:italic> during initiation of bidirectional chromosome replication in <jats:italic>E. coli.</jats:italic></jats:p>