Mutagenesis identifies the critical regions and amino acid residues of suid herpesvirus 1 DNA-binding protein required for DNA binding and strand invasion

Herpesviral DNA-binding protein (DBP) is a unique protein involved in viral DNA replication and genomic recombination. It binds and stabilizes the single-stranded DNA. It also forms the D-loops and promotes the strand invasion. To identify the functional regions and amino acid residues required for DNA binding and D-loop formation, we characterized several DBP mutants of suid herpesvirus 1 (SuHV-1). Acetic anhydride modification assay showed that lysine residues were critical for DNA binding and D-loop formation. Replacement of highly conserved lysine residues with alanine revealed that Lys-756 and Lys-970 were critical for DNA binding, while Lys-161 participated in DNA binding and D-loop formation. Analysis of nested deleted mutants showed that N-terminal 201 amino acid residues and C-terminal 305 amino acid residues were required for D-loop formation and DNA binding, respectively. In conclusion, these findings suggested that SuHV-1 DBP contained critical regions for DNA binding and D-loop formation, and Lys-161, Lys-756, and Lys-970 were required for DNA binding or D-loop formation.