Unlimited Cooperativity of Betatectivirus SSB, a Novel DNA Binding Protein Related to an Atypical Group of SSBs From Protein-Primed Replicating Bacterial Viruses

Abstract

Bam35 and related betatectiviruses are tail-less bacteriophages that prey on members of the Bacillus cereus group. These temperate viruses replicate their linear genome by a protein-primed mechanism. In this work, we have identified and characterized the product of the viral ORF2 as a single-stranded DNA binding protein (hereafter B35SSB). B35SSB binds ssDNA with great preference over dsDNA or RNA in a sequence-independent, highly cooperative manner that results in a non-specific stimulation of DNA replication. We have also identified several aromatic and basic residues, involved in base-stacking and electrostatic interactions, respectively, that are required for effective protein–ssDNA interaction. Although SSBs are essential for DNA replication in all domains of life as well as many viruses, they are very diverse proteins. However, most SSBs share a common structural domain, named OB-fold. Protein-primed viruses could constitute an exception, as no OB-fold DNA binding protein has been reported. Based on databases searches as well as phylogenetic and structural analyses, we showed that B35SSB belongs to a novel and independent group of SSBs. This group contains proteins encoded by protein-primed viral genomes from unrelated viruses, spanning betatectiviruses and Φ29 and close podoviruses, and they share a conserved pattern of secondary structure. Sensitive searches and structural predictions indicate that B35SSB contains a conserved domain resembling a divergent OB-fold, which would constitute the first occurrence of an OB-fold-like domain in a protein-primed genome.