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.
Wu
Shih-Lu
SL
Department of Biochemistry, China Medical University, Taichung 40402, Taiwan.
Li
Chia-Cheng
CC
Ho
Tin-Yun
TY
Hsiang
Chien-Yun
CY
D000595:Amino Acid Sequence; D004277:DNA, Single-Stranded / Q000378:metabolism*; D004279:DNA, Viral / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D011558:Herpesvirus 1, Suid / Q000235:genetics*; D008239:Lysine / Q000235:genetics; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*
D004277:DNA, Single-Stranded; D004279:DNA, Viral; D004268:DNA-Binding Proteins; D011994:Recombinant Proteins; D014764:Viral Proteins; D008239:Lysine