Initiation of chromosomal replication requires dynamic nucleoprotein complexes. In most eubacteria, the originoriCcontains multiple DnaA box sequences to which the ubiquitous DnaA initiators bind. InEscherichia coli oriC, DnaA boxes sustain construction of higher-order complexesviaDnaA–DnaA interactions, promoting the unwinding of the DNA unwinding element (DUE) withinoriCand concomitantly binding the single-stranded (ss) DUE to install replication machinery. Despite the significant sequence homologies among DnaA proteins,oriCsequences are highly diverse. The present study investigated the design oforiC(tma-oriC) fromThermotoga maritima, an evolutionarily ancient eubacterium. The minimaltma-oriCsequence includes a DUE and a flanking region containing five DnaA boxes recognized by the cognate DnaA (tmaDnaA). This DUE was comprised of two distinct functional modules, an unwinding module and atmaDnaA-binding module. Three direct repeats of the trinucleotide TAG within DUE were essential for both unwinding and ssDUE binding bytmaDnaA complexes constructed on the DnaA boxes. Its surrounding AT-rich sequences stimulated only duplex unwinding. Moreover, head-to-tail oligomers of ATP-boundtmaDnaA were constructed withintma-oriC, irrespective of the directions of the DnaA boxes. This binding mode was considered to be induced by flexible swiveling of DnaA domains III and IV, which were responsible for DnaA–DnaA interactions and DnaA box binding, respectively. Phasing of specifictmaDnaA boxes intma-oriCwas also responsible for unwinding. These findings indicate that a ssDUE recruitment mechanism was responsible for unwinding and would enhance understanding of the fundamental molecular nature of the origin sequences present in evolutionarily divergent bacteria.