The atomic structure of the HuH endonuclease domain of HBoV1 NS1 closely resembles the nickase structures encoded by other parvoviruses and by more-distant RCR replicons. This structure also mediates site-specific duplex DNA-recognition, which allows NS1 to bind site-specifically to viral replication origins positioned at each end of its genome (derived from the sequences of the viral hairpin telomeres). Origin recognition, which for some parvoviruses must be enhanced by the binding of additional cellular cofactors, leads to strand- and site-specific nicking of viral duplex DNA replication intermediates, processes that require ATP for tight binding and subsequent nicking. The NS1 protein remains covalently linked to the 5 end of nicked DNA, while the new 3-hydroxyl group is able to prime synthesis of additional linear sequences. Replication of the genome is thought to be mediated by DNA repair DNA polymerases. This process involves the single strand-binding protein replication protein A and NS1. In this process NS1 acts as an ATP powered helicase to resolve terminal hairpin structures of the viral genome.
