A conserved mechanism for replication origin recognition and binding in archaea

Alan I Majerník , James Pj Chong (1)

1 Department of Biology [York]

To date, methanogens are the only group within the Archaea where firing DNA replication origins have not been demonstrated in vivo. Here we show that a previously identified cluster of Origin Recognition Box (ORB) sequences do indeed function as an origin of replication in vivo in the archaeon Methanothermobacter thermautotrophicus. Although the consensus sequence of ORBs in M. thermautotrophicus is somewhat conserved when compared to ORB sequences in other archaea, the Cdc6-1 protein from M. thermautotrophicus (MthCdc6-1) displays sequence-specific binding that is selective for the MthORB sequence and does not recognize ORBs from other archaeal species. Stabilization of in vitro MthORB DNA binding by MthCdc6-1 requires additional conserved sequences 3´ to those originally described for M. thermautotrophicus. By testing synthetic sequences bearing mutations in the MthORB consensus sequence, we show that Cdc6-ORB binding is critically dependent on the presence of an invariant guanine found in all archaeal ORB sequences. Mutation of a universally conserved arginine in the recognition helix of the winged helix domain of archaeal Cdc6-1 shows that specific origin sequence recognition is dependant on the interaction of this arginine with the invariant guanine. Recognition of a mutated origin sequence can be achieved by mutation of the conserved arginine to a lysine or glutamine. Thus, despite a number of differences in protein and DNA sequences between species, the mechanism of origin recognition and binding appears to be conserved throughout the archaea.

Mots clés: Life Sciences

Alan I Majerník, James Pj Chong. A conserved mechanism for replication origin recognition and binding in archaea. Biochemical Journal, 2007, 409 (2), pp.511-518. ⟨10.1042/BJ20070213⟩. ⟨hal-00478756⟩