The RecJ2 Protein in the Thermophilic Archaeon Thermoplasma acidophilum Is a 3′ 5′ Exonuclease and Associates with a DNA Replication Complex [Microbiology]

March 16th, 2017 by Hiromi Ogino, Sonoko Ishino, Daisuke Kohda, Yoshizumi Ishino

RecJ/cell division cycle 45 (Cdc45) proteins are widely conserved in the three domains of life, i.e., in Bacteria, Eukarya and Archaea. Bacterial RecJ is a 5′ 3′ exonuclease and functions in DNA repair pathways, while using its 5′ 3′ exonuclease activity. Eukaryotic Cdc45 has no identified enzymatic activity, but participates in the CMG complex so named because it is composed of Cdc45, minichromosome maintenance protein complex (MCM) proteins 2-7, and GINS complex proteins (Sld5, Psf11 to 3). Eukaryotic Cdc45 and bacterial/archaeal RecJ share similar amino acid sequences and are considered functional counterparts. In Archaea, a RecJ homolog in Thermococcus kodakarensis was shown to associate with GINS and accelerate its nuclease activity and was therefore designated GAN (GINS-associated nuclease); however, to date, no archaeal RecJ MCM GINS complex has been isolated. The thermophilic archaeon Thermoplasma acidophilum has two RecJ like proteins, designated TaRecJ1 and TaRecJ2. TaRecJ1 exhibited DNA-specific 5′ 3′exonuclease activity, while TaRecJ2 had 3′ 5′ exonuclease activity and preferred RNA over DNA. TaRecJ2, but not TaRecJ1, formed a stable complex with TaGINS in a 2:1 molar ratio. Furthermore, the TaRecJ2-TaGINS complex stimulated activity of TaMCM helicase in vitro, and the TaRecJ2-TaMCM-TaGINS complex was also observed in vivo. However, TaRecJ2 did not interact with TaMCM directly and was not required for the helicase activation in vitro. These findings suggest that the function of archaeal RecJ in DNA replication evolved divergently from Cdc45 despite conservation of the CMG-like complex formation between Archaea and Eukarya.
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