Substrate specificity and action mechanism of the HerA-NurA nuclease from the hyperthermophilic archaeon Thermococcus kodakarensis

ABSTRACT The HerA-NurA complex reportedly functions in DNA end resection in archaea. End resection is important to start homologous recombination by forming a single-stranded DNA region with an overhanging 3′-end, which invades double-stranded DNA (dsDNA) with a homologous sequence to form a D-loop. Here, we studied the structure and functions of HerA-NurA from the hyperthermophilic archaeon, Thermococcus kodakarensis. Our analyses demonstrated that NurA is a non-directional and single-stranded specific nuclease, but the HerA-NurA complex cleaves both strands of dsDNA in an exonucleolytic manner, regardless of the structure of the DNA end. The 3D structures of HerA-NurA and its complex with dsDNA revealed the detailed molecular mechanisms of these nuclease reactions. These results suggest that HerA-NurA may not be involved in the end resection process but instead performs other functions, such as exerting an antiviral function by degrading the dsDNA of foreign viruses, similar to recent studies in bacteria.IMPORTANCETo understand the specific function of the HerA-NurA complex, which is believed to function in the end resection process to create a 3′-overhanging DNA for the following strand invasion in homologous recombination, we performed biochemical and structural analyses of this complex from a hyperthermophilic archaeon, Thermococcus kodakarensis, inhabiting a harsh environment where DNA is easily damaged. We found that the HerA-NurA complex cleaves both strands of double-stranded DNA in an exonucleolytic manner, regardless of the structure of the DNA end. Our structural analysis revealed the detailed characteristics of the nuclease activity exhibited by the HerA-NurA complex. Based on the presented information, it is unlikely that the HerA-NurA complex directly functions in end resection, but rather is involved in other functions, possibly in defense against viral infections.