Bioinformatic and genetic characterization of three genes localized adjacent to the major replication origin of Haloferax volcanii

In haloarchaea, a cluster of three genes is localized directly adjacent to the major replication origin, and, hence, the encoded proteins were annotated as 'origin-associated proteins' (Oap). However, prior to this study, no experimental data were available for these conserved hypothetical proteins. Bioinformatic analyses were performed, which unraveled, 1) that the amino acid composition of all three proteins deviate from the average, 2) that OapA is a GTP-binding protein, 3) that OapC has an N-terminal zinc-finger motif, and 4) that the sequences of OapA and OapB are highly conserved while OapC conservation is restricted to short terminal regions. Surprisingly, transcript analyses revealed a complex expression pattern of the oap genes, despite their close proximity. Based on the high degree of conservation in haloarchaea it could be expected that one or more of the oap genes might be essential. However, in frame deletion mutants of all three genes could be readily generated, were viable, and had no growth phenotype. In addition, quantification of the chromsome copy numbers revealed no significant differences between the wild-type and the three mutants. In summary, experimental evidence is inconsistent with Oap proteins being essential for or involved in key steps of DNA replication.